RNA surveillance among curated proteins

ABSTRACT

Computational methods for systematically characterizing putative protein isoforms as apparent targets of nonsense-mediated decay (NMD) comprise: (a) identifying a dataset of target putative protein isoform sequences for characterization; (b) identifying from an mRNA dataset corresponding mRNA sequences representing transcripts encoding the protein isoforms; (c) determining corresponding gene intron-exon structures by mapping the mRNA sequences to corresponding genomic sequences; and (d) determining if the transcripts are apparent targets of NMD. Methods for regulating the expression of a gene encoding a protein isoform characterized as an apparent target of NMD comprise biasing expression of the isoform by modulating transcript splicing or modulating NMD activity.

This invention was made with Government support under Grant Nos.732-HG000747 and 1-K22-HG00056 awarded by the National Institutes ofHealth. The Government has certain rights in this invention.

INTRODUCTION

1. Field of the Invention

The field of the invention is computational and molecular methodsrelating to gene regulation by alternative splicing andnonsense-mediated decay.

2. Background of the Invention

Alternative splicing is an intensely studied cellular phenomenon wherebycells can produce multiple versions of many of the genes encoded intheir genomes. Current estimates are that more than one-half of allhuman genes undergo alternative splicing. We recently disclosed thatmany of these alternative transcripts are targets of a degradationpathway known as nonsense-mediated mRNA decay (NMD) (ref. 1, and U.S.patent application Ser. No. 10/159,997). Here we disclose ourdevelopment of a computational protocol for identifying likely NMDtargeted isoforms from amongst the well-characterized alternativeisoforms in protein databases. For example, applying our protocol to theSwiss-Prot database, we characterized 177 human alternative isoforms astargets of NMD. Amongst these are isoforms of Calpain-10, CLK1, CLK2,CLK3, and LARD/TNFRSF12. We also disclose a protocol for characterizingpolymorphisms as generative of NMD targeted isoforms.

SUMMARY OF THE INVENTION

We extend, adapt and apply our prior invention to analysis of proteinisoform datasets for putative isoforms that are encoded by transcriptsthat are targets of NMD. The invention provides methods forcharacterizing purported protein isoforms, and for using the resultantcharacterization to guide use of the proteins, and particularly tomodulate predetermined isoform expression. The invention providescharacterization protocols, as well as output databases comprising listsof isoforms subject to NMD. The invention also provides protein-specificmethods, such as NMD-based methods for regulating the expression of eachidentified protein. The invention includes subsets of the identifiedprotein isoforms, particularly subsets which exclude any isoformspreviously suggested by others to be NMD targets.

In particular embodiments, the invention provides computational methodsfor characterizing a putative protein isoform as an apparent target ofNMD, comprising the steps of:

(a) identifying a target putative protein isoform sequence forcharacterization;

(b) identifying one or more corresponding mRNA sequences representing atranscript encoding the protein isoform;

(c) determining corresponding gene intron-exon structure by mapping themRNA sequences to one or more corresponding genomic sequences;

(d) determining if the transcript is an apparent target of NMD.

In particular embodiments, the invention provides computational methodsfor systematically characterizing putative protein isoforms as apparenttargets of NMD, comprising the steps of:

(a) identifying a dataset of target putative protein isoform sequencesfor characterization;

(b) identifying from an mRNA dataset corresponding mRNA sequencesrepresenting transcripts encoding the protein isoforms;

(c) determining corresponding gene intron-exon structures by mapping themRNA sequences to corresponding genomic sequences; and

(d) determining if the transcripts are apparent targets of NMD.

In particular embodiments, the invention provides computational methodsfor systematically characterizing putative protein isoforms as apparenttargets of NMD, comprising the steps of:

(a) identifying a dataset of target putative protein isoform sequencesfor characterization;

(b) identifying corresponding gene intron-exon structure by mappingcorresponding mRNA sequences of an mRNA sequence dataset to genomicsequences of a genomic DNA sequence dataset;

(c) screening the isoform dataset for a subset of isoforms encoded bytranscripts comprising alternate splices which introduce a stop codonmore than 50 bp upstream of the final exon-exon splice junction; and

(d) classifying the subset isoforms as being encoded by transcriptscomprising premature stop codons and as targets of NMD.

In particular embodiments, the invention further comprises the step ofcorrelating the isoforms with relative expression in different celltypes, particularly cancer cells and non-cancer cells.

The invention also provides methods for regulating the expression of agene encoding a protein isoform of Table 1, said method comprising thestep of biasing expression of the isoform by modulating transcriptsplicing or modulating NMD activity, particularly wherein the isoformexpression is biased with an inhibitor selected from the groupconsisting of: an aminoglycoside drug like gentamycin which causestranslational readthrough; a dominant negative hUpf1, expression ofwhich inhibits NMD; an hUpf1-targeted RNAi which inhibits NMD; anantisense exon or a specific splicing factor, expression of whichinhibits NMD; and a splice-selection altering amount of dimethylsulfoxide which inhibits NMD.

The invention also provides datasets of putative protein isoformscharacterized as apparent targets of NMD by a subject method,particularly wherein the datasets comprise a subset of the isoforms ofTable 1, particularly isoforms of Calpain-10, CLK1, CLK2, CLK3, andLARD/TNFRSF12. These datasets may be in any convenient form, includingstored in or on an electronic medium.

DETAILED DESCRIPTION OF PARTICULAR EMBODIMENTS OF THE INVENTION

The following descriptions of particular embodiments and examples areoffered by way of illustration and not by way of limitation

We disclose that about 10% of the alternatively spliced entries in theSWIS-PROT protein database have a premature termination codon, makingthem candidates for nonsense-mediated decay. Our findings indicate thatmany purported alternatively spliced protein forms naturally exist inlower abundance than previously thought, and in many cases NMD reducestheir abundance to undetectable and insignificant levels. We provide alist of several hundred such purported isoforms, and detailed exemplaryanalyses of three of them. Our findings provide an unappreciatedapproach—selective NMD targeting—for regulating the expression of theseproteins.

I. Computational method for identifying likely NMD-targets in Swiss-Protand other highly curated databases. Many curated biological sequencedatabases support annotation of alternative splicing. Because thecloning and characterization of many genes predates our currentunderstanding of NMD in vertebrates, many of the alternative isoformsdescribed in these databases are unrecognized targets of NMD. A seriesof recent experiments from several labs has outlined the mechanisticdetails of vertebrate NMD. The model of NMD demonstrated by theseexperiments combined with the following computational analysis of mRNAtranscripts allows us to predict which isoforms are likely targets forNMD. Knowing that a transcript is degraded by NMD is criticallyimportant for understanding its function. It is largely assumed thatgenes that are expressed as mRNA go on to be expressed as protein.However, when NMD intervenes, little or no protein product is made andthe gene is effectively off.

Our method for discovering which alternative isoforms are likely NMDtargets from a database of isoform sequences is as follows. First, weidentify the transcript sequence of each isoform from a relevantdatabase. This database may or may not be separate from the databasebeing screened. Then, using complete genomic sequence, we use SPIDEY ora similar transcript-to-genome alignment program to determine whereintrons have been spliced out of each transcript. A comparison of theposition of the termination codon to the position of introns thenreveals if the transcript is likely to be recognized and degraded byNMD. If the termination codon is more than 50 nucleotides upstream ofthe final intron position, then the transcript is an apparent target ofNMD.

II. Identification of 177 human alternative isoforms from Swiss-Protthat are likely targets of NMD including Calpain-10, CLK1, CLK2, andCLK3, and LARD/TNFRSF12. Using the above protocol, we screened the humanalternative isoforms of the Swiss-Prot database. This screen identified130 alternatively spliced genes that generate 177 isoform transcriptsthat are likely targets of NMD.

Further investigation of several of these revealed that they arebona-fide targets of NMD. Calpain-10, for example, is a ubiquitouslyexpressed protease that is alternatively spliced to produce eightisoforms. A literature report describes four of these isoforms as “lessabundant”. These same four isoforms were identified in our screen to belikely NMD targets.

We also identified as likely NMD targets isoforms of the human spliceregulators CLK1, CLK2, and CLK3. CLKs, also known as LAMMER kinases,have been shown to regulate alternative splicing by phosphorylating andthereby activating SR proteins. Alignment of the CLK1, CLK2, and CLK3human paralogs reveals that all three share a conserved splicing patterncomprised of a full-length isoform and a shorter, premature terminationcodon (PTC)-containing isoform generated by skipping exon 4. EST datashow this alternative splicing pattern is evolutionarily conservedthrough mice and sea squirts. A prior investigation of the relativeexpression of the CLK1 splice variants shows that the PTC-containingisoform is “up-regulated” relative to the non-PTC isoform followingtreatment with cycloheximide, a potent NMD inhibitor. While thisup-regulation was previously attributed to the cellular stress response,our results indicate alleviation of NMD is responsible for the higherabundance of the PTC form.

Intriguingly, CLK1 has also been shown to affect its own splicing. Thepresence of high levels of CLK1 protein favors generation of thePTC-containing mRNA isoform that we expect to be targeted by NMD. Thismay set up an auto-regulatory feedback loop analogous to that seen forSC35, which auto-regulates its own expression via alternativesplicing-induced NMD.

We also identified several isoforms of the death domain-containingreceptor TNFRSF12/LARD/DR3/Apo3 as likely NMD targets. Deathdomain-containing receptor proteins regulate the balance betweenlymphocyte proliferation and apoptosis. Resting lymphocytes expressseveral short TNFRSF12 isoforms of unclear function. In contrast,activated lymphocytes produce only the full-length TNFRSF12, whichinduces apoptosis. We found that the five short TNFRSF12 mRNA isoformsexpressed in resting lymphocytes all had PTCs. This precise correlationbetween mRNA isoform expression and NMD-candidacy indicates that theshort variants are simply degraded, while the full-length version is theonly one translated normally to yield protein. Alternative splicingcould thus act as a switch, regulating whether protein is expressed andthus whether the cell undergoes apoptosis.

III. Computational protocol for screening polymorphism libraries todetect NMD-targeted polymorphisms. There are several large public andproprietary databases of naturally occurring human single-nucleotidepolymorphisms (SNPs). SNPs have been used for a variety of purposes.Perhaps the most common use of SNP data is in disease-linkage studies.When a specific SNP is associated with a specific disease, it can givevaluable clues to the role of the gene in which the SNP occurs. Usingour protocol for determining gene structures, we can make the additionalassessment of whether any given SNP will likely generate anNMD-candidate isoform. In these cases, disease pathology will likely bedue to haplo-insufficiency rather than gain-of-function.

IV. Control of protein expression by the manipulation of splicingfactors. We demonstrate that genes can be generally and predictablyengineered to undergo alternative splicing, so that multiple mRNAisoforms are generated. Engineered alternative splicing can be regulatedin any of the ways that alternative splicing is regulated in vivo:addition or subtraction of known alternative splicing factors, changingconcentration of basal splicing factors, etc. One or more of theisoforms can be engineered to contain premature termination codonscausing these mRNAs to be selectively degraded. In this way, underprescribed conditions, engineered genes can be tightly regulated.Accordingly, the novel isoforms disclosed herein provide myriadreagents, including sites, splice junctions and splice environments,with pre-determinable effects on target gene expression regulation.While precise effects are best confirmed empirically, native expressionpatterns of the natural target gene provide preliminary guidance forpredetermining heterologous expression regulation. A particularadvantage of this post-transcriptional regulation is that it can makeuse of endogenous splicing factors to control expression, making themethod particularly well-suited for in vivo applications. Hence, thismethod is useful for studying the function of specific genes andproteins in cell culture, and for generating transgenic animals in whichexpression of exogenous genetic material is limited to certain celltypes based on their splicing environment.

Accordingly, the invention provides engineering target genes to undergoalternative splicing, so that multiple mRNA isoforms, including one ormore NMD-targeted isoforms, are generated, and thereby providingpredetermined NMD-mediated, postranscriptional expression regulation ofthe gene. In a particular embodiment, the invention provides a methodfor regulating gene expression comprising the steps of:

genetically engineering in a target gene a change in the native patternof splice junctions to provide at least a predetermined first,non-natural NMD-targeted splice form and a predetermined second,non-NMD-targeted splice form, wherein expression of the gene isregulated by the relative expression of the first and second spliceforms; and optionally,

detecting or inferring the relative expression of the first and secondsplice forms; and optionally,

detecting or inferring a change in expression of corresponding proteinisoforms.

Protocols for manipulating splicing and/or NMD are known in the art. Forexample, aminoglycoside drugs, like gentamicin, cause translationalreadthrough, and therefore inhibit NMD (e.g. Mankin and Liebman, NatGenet. 1999 September; 23(1):8-10; Burke and Mogg, Nucleic Acids Res.1985 Sep. 11; 13(17):6265-72); expression of a dominant negative versionof hUpf1 inhibits NMD (e.g. Sun et al., Proc Natl Acad Sci USA. 1998Aug. 18; 95(17):10009-14); RNAi directed against hUpf1 inhibits NMD(e.g. Mendell et al., Science 2002 Oct. 11; 298(5592):419-22);expressing antisense exons can prevent expression of NMD-targetedisoforms (e.g. Mann et al., Proc Natl Acad Sci USA. 2001 Jan. 2;98(1):42-7); expressing specific splicing factors can prevent expressionof NMD-targeted isoforms (e.g. Caceres and Kornblihtt, Trends inGenetics 2002 Apr. 18(4): 186-193); and chemical reagents like DMSO canprevent expression of NMD-targeted isoforms (e.g. Bolduc et al., J BiolChem. 2001 May 18; 276(20):17597-602)

V. NMD targeted isoforms in diseased cells. The invention provides forthe application of any of the foregoing experimental or computationalmethods in the diagnosis of disease states, such as cancer, and in thedevelopment of diagnoses and therapies based on NMD-mediated geneexpression regulation.

EXAMPLES

Alternative pre-mRNA splicing endows genes with the potential to producea menagerie of protein products. We recently found that 35% of reliableEST-inferred human alternative mRNA isoforms containpremature-termination codons (PTCs), rendering them candidate targets ofa surveillance pathway known as nonsense-mediated mRNA decay (NMD) (1).This finding indicates that many alternative mRNA isoforms are nottranslated into functional protein but are instead targeted fordegradation. Because the cloning and characterization of many genespredates an appreciation of the pervasiveness of NMD, we hypothesizedthat curated databases may inadvertently contain isoforms that are NMDtargets. Here we report that 9.6% of the alternatively spliced humanentries in SWISS-PROT that were amenable to analysis have at least onePTC-containing splice variant, making them apparent targets of NMD(Table 1).

We assembled 4556 human protein isoform sequences from 1636alternatively-spliced human entries in the SWISS-PROT database v.41(2).We next located corresponding mRNA sequences in the REFSEQ and GENBANKbioinformatics databases and determined gene structures using SPIDEY(3)and the NCBI human genome build 30. We discarded SWISS-PROT isoformsequences for which we could not locate the corresponding mRNA sequenceor assemble a suitable SPIDEY alignment. The “50-nucleotide rule” forPTCs and NAND (ref. 4 and references therein) was checked for each mRNAby comparing the position of the termination codon relative to theintrons (Example Supplement 1). Of the 2523 isoform sequences from 1348SWISS-PROT entries that passed quality filters, 177 splice variantisoforms (7.0%) from 130 entries (9.6%) were found to contain a PTC.

For several of these genes, experimental reports include data consistentwith NMD action, though this possibility has been almost universallyoverlooked. For example, literature reports regarding Calpain-10describe isoform abundances that correlate perfectly with our NMDprediction (Example Supplement 2). Horikawa et al. describe four ofCalpain-10's eight splice variants as being expressed in “lowabundance.” (5) It is these same four isoforms that were found tocontain PTCs, implicating NMD as the culprit behind the experimentalobservation.

Alternative isoforms of human CLK1, CLK2, and CLK3 were also identifiedin our analysis as containing PTCs. All three share a conserved splicingpattern comprised of a full-length isoform and a shorter, PTC-containingisoform generated by skipping exon 4(6). EST data show this alternativesplicing pattern is evolutionarily conserved through mice and seasquirts (Example Supplement 3). A prior investigation of the relativeexpression of the CLK1 splice variants shows that the PTC-containingisoform is “up-regulated” relative to the non-PTC isoform followingtreatment with cycloheximide, a potent NMD inhibitor (7). Menegay et al.attributed this up-regulation to the cellular stress response (7); ourresults indicate alleviation of NMD is responsible.

CLKs are alternative splicing regulators and CLK1 has also been shown toaffect its own splicing (8). The presence of high levels of CLK1 proteinfavors generation of the PTC-containing isoform creating a feedback loopanalogous to that of SC35 (9), which auto-regulates its own expressionvia alternative splicing-induced NMD.

Literature reports describing the expression of TNFRSF12/LARD/DR3/Apo3isoforms contain data consistent with NMD-associated regulation. Ouranalysis found six TNFRSF12 protein isoforms with PTCs. Prior tolymphocyte activation, only those isoforms that we found to contain PTCsare expressed (10). Following PHA-mediated lymphocyte activation,expression of these isoforms ceases in favor of the major, full-lengthisoform (10). This shift in splicing hints that alternativesplicing-induced NMD may play a role in regulating lymphocyte apoptosis(Example Supplement 4). Prior to lymphocyte activation, the regulatedsplicing of unproductive isoforms, degraded by NMD, would hold inabeyance apoptosis which is induced by splicing the productive isoform.

Example Supplement 1 Methods

(a) Extraction and assembly of alternatively spliced human proteinisoforms from SWISS-PROT v41. We analyzed each of the 2292 SWISS-PROThuman entries containing a VARSPLIC line in its feature table. VARSPLICline information was used to assemble protein isoform sequence for 4556isoforms from 1636 SWISS-PROT entries. 656 entries could not be analyzeddue to ambiguous VARSPLIC annotation.

(b) Identification of corresponding mRNA/cDNA sequences. AlthoughSWISS-PROT contains cross-references to mRNA/cDNA sequences for majorprotein isoforms, many alternative isoforms are not cross-referenced. Tofind the correct cDNA/mRNA isoform sequence for each SWISS-PROT proteinisoform, we used BLAST version 2.2.4 to align each protein isoformsequence to translated mRNA/cDNA sequences from the REFSEQ and GENBANKbioinformatics databases of Mar. 22, 2003. In these alignments, >99%identity over the full length of the SWISS-PROT isoform was required. Incases of multiple matches, we selected 100% identical matches over 99%identical matches and REFSEQ matches over GENBANK matches. ForSWISS-PROT isoforms matching multiple entries from the same database atthe same percent identity, the match associated with the longest mRNAsequence was chosen. After applying these rules, 2787 alternativelyspliced human Swiss-Prot protein isoforms were associated with acorresponding cDNA/mRNA sequences from either REFSEQ or GENBANK.

(c) Retrieving coding sequences and genomic loci. We used LOCUSLINKbioinformatics database to map each cDNA/mRNA sequence to the correcthuman genomic contig sequence from the NCBI human genome build 30. TheCDS feature of each REFSEQ or GENBANK bioinformatics database record wasused to identify the location of the termination codon.

(d) Assessing NMD candidacy. The SPIDEY mRNA to genomic alignmentprogram was used to determine the location of introns for each cDNA/mRNAisoform sequence. SPIDEY takes as input a cDNA/mRNA sequence and thecorresponding genomic sequence and generates an alignment that revealsthe location of introns. We compared the intron positions to theposition of the termination codon for each cDNA/mRNA isoform sequence.If the termination codon was found to be more than 50 nucleotidesupstream of the final intron, it was deemed a premature-terminationcodon and the transcript deemed an apparent target for NMD. This was thecase for 7.0% of the isoforms (177 of 2523) from 9.6% of the SWISS-PROTentries (130 of 1348).

Example Supplement 2

Expression levels of Calpain-10 isoforms are consistent with NMDprediction. Calpain-10 (Q9HC93), a ubiquitously expressed protease, isalternatively spliced to produce eight isoforms. An initial report fromHorikawa and co-workers showed that several of the isoforms areexpressed in low abundance. This set of isoforms correlates perfectlywith the set that we determined to be NMD-candidates. This study alsoreported that a SNP in Calpain-10 is strongly linked to susceptibilityto Type-II diabetes in several populations. This mutation lies inintronic sequence and may affect the alternative splicing of Calpain-10.

Example Supplement 3

Splicing to generate premature termination codon is evolutionarilyconserved in CLKs. The cdc-like kinases (CLKs) control alternativesplicing by phosphorylating SR-proteins. CLKs are alternatively spliced,themselves, in a pattern that is conserved from human through mouse andsea-squirt.

(a) Conservation across three human paralogs. Our screen of Swiss-Protrevealed that human CLK1, CLK2, and CLK3 generate premature terminationcodon (PTC) containing isoforms. Using SPIDEY, GFF2PS, and customscripts, we generated gene structures that show both the full-lengthisoforms and the PTC containing isoform. The splicing pattern thatgenerates the putative NMD target isoforms, skipping exon 4, isconserved in each. We also generated global alignments betweencorresponding exons and introns using ALIGN. The introns flanking thealternative exon are amongst the most conserved.

(b) Conservation across orthologs in human, mouse, and sea-squirt. CLKswere identified in mouse through existing annotation and in thepredicted genes of Ciona Intestinalis (sea-squirt) through HMMER searchusing an HMM constructed with annotated CLKs from a variety oforganisms. Pairwise alignment revealed the sea-squirt CLK to be mostsimilar to CLK2. We then identified ESTs from dbEST corresponding to thethree mouse CLKs and to the sea-squirt CLK. Analysis of these ESTsrevealed that the PTC-generating alternative splicing pattern wasconserved in each. Corresponding exons and introns were also aligned asbefore. However, the human CLK2 gene has 13 exons whereas sea-squirt CLKgene has 11. To ensure that we aligned homologous regions, we generatedclustalw and t_coffee multiple sequence alignments of annotated CLKsfrom several organisms and used these alignments to guide our exon andintron alignments. This analysis clearly indicated a single extra intronin sea squirt relative to human and three extra introns in humanrelative to sea-squirt. As in the paralog comparison, a high degree ofconservation is present in the introns flanking the alternative exon.

Example Supplement 4

TNFRSF12/LARD/DR3/Apo3 Splicing Pattern and Expression Levels. TNFRSF12is a death domain-containing member of the nerve growth factor receptor(NGFR) family of proteins that is found almost exclusively inlymphocytes. The term “death domain” refers to a conserved intracellularregion found in receptors like Fas and TNFR-1 that is capable ofinducing apoptosis while in the presence of a particular ligand (in thiscase, FasL and TNF1 respectively). Death domain-containing receptorsplay a crucial role in maintaining the balance between lymphocyteproliferation and apoptosis in vivo. TNFRSF12 is alternatively splicedto produce at least 11 isoforms, though only the major splice variant(isoform 1) contains the death domain and is capable of inducingapoptosis. Six TNFRSF12 isoforms were found in our analysis ofSWISS-PROT to contain PTCs, rendering them potential targets of (NMD.The splicing distribution of TNFRSF12 isoforms has been shown to changeupon lymphocyte activation, indicating alternative splicing may be acontrol point regulating lymphocyte proliferation.

(a) Screaton et al. (10) showed that, prior to lymphocyte activation,only PTC-containing TNFRSF12 isoforms are expressed (isoforms 2, 3, 4,and 5). Primary blood lymphocytes treated with an activating agent werefound to instead express the major, apoptosis-inducing splice variant(isoform 1) almost exclusively. The correlation between PTC-containingisoform expression and lymphocyte activation indicates alternativesplicing-induced NMD plays a role in regulating lymphocyteproliferation. By expressing only PTC-containing isoforms prior toactivation, programmed cell death would be held in abeyance. Onceactivated, lymphocytes could alter splicing patterns to favor expressionof full-length TNFRSF12 isoform 1, inducing apoptosis.

(b) TNFRSF12 isoforms 2, 3, 4, and 5 were found in our analysis ofSWISS-PROT to contain premature termination codons (PTCs), renderingthem apparent targets of NMD.

PARENTHETICAL REFERENCES

-   1. Lewis, B. P., Green, R. E. & Brenner, S. E. Evidence for the    widespread coupling of alternative splicing and nonsense-mediated    mRNA decay in humans. Proc Natl Acad Sci USA 100, 189-92 (2003).-   2. Boeckmann, B. et al. The SWISS-PROT protein knowledgebase and its    supplement TrEMBL in 2003. Nucleic Acids Res 31, 365-70 (2003).-   3. Wheelan, S. J., Church, D. M. & Ostell, J. M. Spidey: a tool for    mRNA-to-genomic alignments. Genome Res 11, 1952-7 (2001).-   4. Maquat, L. E. Nonsense-mediated mRNA decay. Curr Biol 12, R 196-7    (2002).-   5. Horikawa, Y. et al. Genetic variation in the gene encoding    Calpain-10 is associated with type 2 diabetes mellitus. Nat Genet    26, 163-75 (2000).-   6. Hanes, J., von der Kammer, H., Klaudiny, J. & Scheit, K. H.    Characterization by cDNA cloning of two new human protein kinases.    Evidence by sequence comparison of a new family of mammalian protein    kinases. J Mol Biol 244, 665-72 (1994).-   7. Menegay, H. J., Myers, M. P., Moeslein, F. M. & Landreth, G. E.    Biochemical characterization and localization of the dual    specificity kinase CLK1. J Cell Sci 113 (Pt 18),-   8. Duncan, P. I., Stojdl, D. F., Marius, R. M. & Bell, J. C. In vivo    regulation of alternative pre-mRNA splicing by the Clk1 protein    kinase. Mol Cell Biol 17, 5996-6001 (1997).-   9. Sureau, A., Gattoni, R., Dooghe, Y., Stevenin, J. & Soret, J.    SC35 autoregulates its expression by promoting splicing events that    destabilize its mRNAs. Embo J 20, 1785-96 (2001).-   10. Screaton, G. R. et al. LARD: a new lymphoid-specific death    domain containing receptor regulated by alternative pre-mRNA    splicing. Proc Natl Acad Sci USA 94, 4615-9 (1997).

All publications and patent applications cited in this specification andall references cited therein are herein incorporated by reference as ifeach individual publication or patent application or reference werespecifically and individually indicated to be incorporated by reference.

Although the foregoing invention has been described in some detail byway of illustration and example for purposes of clarity ofunderstanding, it will be readily apparent to those of ordinary skill inthe art in light of the teachings of this invention that certain changesand modifications may be made thereto without departing from the spiritor scope of the appended claims.

TABLE 1 SWISS-PROT Protein isoforms characterized as targets of NMD.ISOFORM NAME ACCESSION PROTEIN DESCRIPTION ISOFORM NAME ACCESSIONPROTEIN DESCRIPTION 3BP2_HUMAN P78314 SH3 domain-binding protein 2HFE_HUMAN Q9HC68 Hereditary ISOFORM SHORT MAJOR hemochromatosis proteinISOFORM precursor 5H4_HUMAN Q13639 5-hydroxytryptamine 4 receptorI17S_HUMAN Q9NRM6 Interleukin-17B receptor MAJOR ISOFORM ISOFORM 2precursor A1A1_HUMAN P05023 Sodium/potassium-transporting ICE2_HUMANP42575 Caspase-2 precursor ISOFORM SHORT ATPase alpha-1 chain precursorISOFORM 1CH-1S ABCD_HUMAN Q9NSE7 Putative ATP-binding cassetteICE8_HUMAN Q9C0K4 Caspase-8 precursor ISOFORM 2 transporter C13 ISOFORM7 ABCD_HUMAN Q9NSE7 Putative ATP-binding cassette ICEA_HUMAN Q92851Caspase-10 precursor ISOFORM 3 transporter C13 ISOFORM B AD11_HUMANO75078 ADAM 11 precursor ICEA_HUMAN Q92851 Caspase-10 precursor ISOFORMSHORT ISOFORM C AD12_HUMAN O43184 ADAM 12 precursor ILF1_HUMAN Q01167Interleukin enhancer- MAJOR ISOFORM ISOFORM 2 binding factor 1AD22_HUMAN Q9P0K1 ADAM 22 precursor IRF7_HUMAN Q92985 Interferonregulatory ISOFORM 2 ISOFORM C factor 7 AKP1_HUMAN Q92667 A kinaseanchor protein 1 IRL1_HUMAN Q01638 Interleukin 1 receptor-like ISOFORM 2mitochondrial precursor ISOFORM C 1 precursor ANPB_HUMAN P20594 Atrialnatriuretic peptide receptor ITP1_HUMAN O14713 Integrin beta-1 bindingISOFORM SHORT B precursor MAJOR protein 1 ISOFORM AS13_HUMAN Q8WXK3Ankyrin repeat and SOCS box ITP1_HUMAN O14713 Integrin beta-1 bindingMAJOR ISOFORM containing protein 13 ISOFORM 2 protein 1 AS13_HUMANQ8WXK3 Ankyrin repeat and SOCS box KC11_HUMAN Q9HCP0 Casein kinase Igamma 1 ISOFORM 2 containing protein 13 ISOFORM 1S isoform ATF3_HUMANP18847 Cyclic-AMP-dependent KLK2_HUMAN P20151 Glandular kallikrein 2ISOFORM 2 transcription factor ATF-3 ISOFORM 3 precursor ATR_HUMANQ9H6X2 Anthrax toxin receptor precursor KLKF_HUMAN Q9H2R5 Kallikrein 15precursor MAJOR ISOFORM ISOFORM 2 B3G7_HUMAN Q9NY97 Beta-13-galactosyltransferase 7 LEF1_HUMAN Q9UJU2 Lymphoid enhancer ISOFORM 2ISOFORM B binding factor 1 BC12_HUMAN Q9HB09 Bcl-2 related proline-richprotein LFA3_HUMAN P19256 Lymphocyte function- ISOFORM 2 ISOFORMassociated antigen 3 SHORT precursor BMP1_HUMAN P13497 Bonemorphogenetic protein 1 LIK1_HUMAN P53667 LIM domain kinase 1 ISOFORMBMP1-4 precursor ISOFORM 3 BMP1_HUMAN P13497 Bone morphogenetic protein1 LSHR_HUMAN P22888 Lutropin- ISOFORM BMP1-5 precursor MAJORchoriogonadotropic ISOFORM hormone receptor precursor BMP1_HUMAN P13497Bone morphogenetic protein 1 LYST_HUMAN Q99698 Lysosomal traffickingISOFORM BMP1-6 precursor MAJOR regulator ISOFORM C343_HUMAN Q9HB55Cytochrome P450 3A43 M2A2_HUMAN P49641 Alpha-mannosidase IIx ISOFORM 4ISOFORM SHORT CA34_HUMAN Q01955 Collagen alpha 3 MADI_HUMAN O95405Mothers against MAJOR ISOFORM ISOFORM 2 decapentaplegic homologinteracting protein CA34_HUMAN Q01955 Collagen alpha 3 MAP2_HUMAN P11137Microtubule-associated ISOFORM 3 MAJOR protein 2 ISOFORM CAL0_HUMANP01258 Calcitonin precursor [Contains: MAP2_HUMAN P11137Microtubule-associated ISOFORM 2 Calcitonin; Katacalcin ISOFORM protein2 MAP2C CANA_HUMAN Q9HC93 Calpain 10 MAP4_HUMAN P27816Microtubule-associated ISOFORM B ISOFORM 2 protein 4 CANA_HUMAN Q9HC93Calpain 10 MAX_HUMAN P25912 Max protein ISOFORM D ISOFORM 3 CANA_HUMANQ9HC93 Calpain 10 MK11_HUMAN Q15759 Mitogen-activated protein ISOFORM EISOFORM kinase 11 BETA-2 CANA_HUMAN Q9HC93 Calpain 10 MLH3_HUMAN Q9UHC1DNA mismatch repair ISOFORM F ISOFORM 2 protein Mlh3 CBS_HUMAN P35520Cystathionine beta-synthase MRP3_HUMAN O15438 Canalicular multispecificMAJOR ISOFORM ISOFORM 3A organic anion transporter 2 CD38_HUMAN P28907ADP-ribosyl cyclase 1 MRP3_HUMAN O15438 Canalicular multispecificISOFORM 2 ISOFORM 3B organic anion transporter 2 CD47_HUMAN Q08722Leukocyte surface antigen CD47 MSRE_HUMAN P21757 Macrophage scavengerISOFORM OA3- precursor ISOFORM 11 receptor types I and II 305 CFLA_HUMANO43618 CASP8 and FADD-like apoptosis MTF2_HUMAN Q9Y483 Metal-responseelement- ISOFORM 9 regulator precursor MAJOR binding transcriptionISOFORM factor 2 CHRD_HUMAN Q9P0Z5 Chordin precursor NK31_HUMAN Q99801Homeobox protein Nkx- ISOFORM 3 MAJOR 31 ISOFORM CHRD_HUMAN Q9P0Z5Chordin precursor NXF5_HUMAN Q9H1B4 Nuclear RNA export ISOFORM 4 MAJORfactor 5 ISOFORM CIQ2_HUMAN O43526 Potassium voltage-gated channelNXF5_HUMAN Q9H1B4 Nuclear RNA export ISOFORM 3 subfamily KQT member 2ISOFORM B factor 5 CIW4_HUMAN Q9NYG8 Potassium channel subfamily KNXF5_HUMAN Q9H1B4 Nuclear RNA export ISOFORM 2 member 4 ISOFORM C factor5 CLK1_HUMAN P49759 Protein kinase CLK1 NXF5_HUMAN Q9H1B4 Nuclear RNAexport ISOFORM SHORT ISOFORM D factor 5 CLK2_HUMAN P49760 Protein kinaseCLK2 NXF5_HUMAN Q9H1B4 Nuclear RNA export ISOFORM SHORT ISOFORM E factor5 CLK3_HUMAN P49761 Protein kinase CLK3 PHMX_HUMAN Q96QS1 Phemx proteinISOFORM 2 ISOFORM 4 CML1_HUMAN Q99788 Chemokine receptor-like 1PHMX_HUMAN Q96QS1 Phemx protein MAJOR ISOFORM ISOFORM 5 COG4_HUMANQ9H9E3 Conserved oligomeric Golgi PML_HUMAN P29590 Probabletranscription ISOFORM 2 complex component 4 ISOFORM PML- factor PML 3BCOLQ_HUMAN Q9UP88 Acetylcholinesterase collagenic PPE1_HUMAN O14829Serine/threonine protein ISOFORM VII tail peptide precursor ISOFORM 2phosphatase with EF- hands-1 CPXM_HUMAN Q96SM3 Potentialcarboxypeptidase X PPT2_HUMAN Q9UMR5 Palmitoyl-protein ISOFORM 2precursor ISOFORM 2 thioesterase 2 precursor CRN1_HUMAN Q9NYD8 Crookedneck-like protein 1 PRD7_HUMAN Q9NQW5 PR-domain zinc finger ISOFORM 4MAJOR protein 7 ISOFORM CRN1_HUMAN Q9NYD8 Crooked neck-like protein 1PSA7_HUMAN O14818 Proteasome subunit alpha ISOFORM 5 ISOFORM 4 type 7CT24_HUMAN Q9BUV8 Protein C20orf24 PSD4_HUMAN P55036 26S proteasome non-ISOFORM 4 ISOFORM ATPase regulatory subunit 4 RPN10E CTGF_HUMAN P29279Connective tissue growth factor PSN1_HUMAN P49768 Presenilin 1 MAJORISOFORM precursor ISOFORM I-374 CU07_HUMAN P57077 Protein C21orf7PTPD_HUMAN P23468 Protein-tyrosine ISOFORM B MAJOR phosphatase deltaISOFORM precursor CU07_HUMAN P57077 Protein C21orf7 R51D_HUMAN O94908DNA repair protein ISOFORM C ISOFORM 2 RAD51 homolog 4 CU18_HUMAN Q9NVD3Protein C21orf18 RBMS_HUMAN Q93062 RNA-binding protein with ISOFORM BMAJOR multiple splicing ISOFORM CU63_HUMAN P58658 Protein C21orf63precursor RED1_HUMAN P78563 Double-stranded RNA- ISOFORM B MAJORspecific editase 1 ISOFORM CU80_HUMAN Q9Y2G5 Protein C21orf80 RHD_HUMANQ9UQ21 Blood group Rh ISOFORM B MAJOR ISOFORM CYB5_HUMAN P00167Cytochrome b5 RHD_HUMAN Q9UQ21 Blood group Rh ISOFORM 2 ISOFORM 3DACA_HUMAN Q9BYJ9 Dermatomyositis associated with RYK_HUMAN P34925Tyrosine-protein kinase ISOFORM 2 cancer putative autoantigen-1 MAJORRYK precursor ISOFORM DFFB_HUMAN O76075 DNA fragmentation factor 40 kDaRYK_HUMAN P34925 Tyrosine-protein kinase ISOFORM BETA subunit ISOFORM 2RYK precursor DFFB_HUMAN O76075 DNA fragmentation factor 40 kDaSCA1_HUMAN O15126 Secretory carrier- ISOFORM DELTA subunit ISOFORM 2associated membrane protein 1 DFFB_HUMAN O76075 DNA fragmentation factor40 kDa SEN7_HUMAN Q9BQF6 Sentrin-specific protease 7 ISOFORM subunitMAJOR GAMMA ISOFORM DJB2_HUMAN P25686 DnaJ homolog subfamily BSFR5_HUMAN Q13243 Splicing factor MAJOR ISOFORM member 2 ISOFORMarginine/serine-rich 5 SRP40-2 DJB2_HUMAN P25686 DnaJ homolog subfamilyB SHX2_HUMAN O60902 Short stature homeobox ISOFORM 3 member 2 MAJORprotein 2 ISOFORM DONS_HUMAN Q9NYP3 Downstream of son gene proteinSNB2_HUMAN Q13425 Beta-2-syntrophin ISOFORM 2 ISOFORM 2 DONS_HUMANQ9NYP3 Downstream of son gene protein SNXD_HUMAN Q9Y5W8 Sorting nexin 13ISOFORM 3 ISOFORM 2 DPP3_HUMAN Q9NY33 Dipeptidyl-peptidase III SON_HUMANQ9UPY0 SON protein ISOFORM 2 ISOFORM C DSCA_HUMAN O60469 Down syndromecell adhesion SON_HUMAN Q9UPY0 SON protein ISOFORM SHORT moleculeprecursor ISOFORM E DTNB_HUMAN O60941 Dystrobrevin beta SUR5_HUMANQ15528 Surfeit locus protein 5 ISOFORM 3 ISOFORM SURF5A EPA3_HUMANP29320 Ephrin type-A receptor 3 T10B_HUMAN O14763 Tumor necrosis factorMAJOR ISOFORM precursor MAJOR receptor superfamily ISOFORM member 10Bprecursor ERAL_HUMAN O75616 GTP-binding protein era homolog T10B_HUMANO14763 Tumor necrosis factor ISOFORM HERA-B ISOFORM receptor superfamilySHORT member 10B precursor ESR2_HUMAN Q9UHD3 Estrogen receptor betaTM31_HUMAN Q9BZY9 Tripartite motif protein 31 ISOFORM 3 ISOFORM BETAF263_HUMAN Q16875 6-phosphofructo-2- TNR6_HUMAN P25445 Tumor necrosisfactor MAJOR ISOFORM kinase/fructose-2 6-biphosphatase 3 ISOFORM 2receptor superfamily member 6 precursor FAFY_HUMAN O00507 Probableubiquitin carboxyl- TNR6_HUMAN P25445 Tumor necrosis factor ISOFORMSHORT terminal hydrolase FAF-Y ISOFORM 3 receptor superfamily member 6precursor FCAR_HUMAN P24071 Immunoglobulin alpha Fc receptor TNR6_HUMANP25445 Tumor necrosis factor ISOFORM B- precursor ISOFORM 4 receptorsuperfamily DELTA-S2 member 6 precursor FCE2_HUMAN P06734 Low affinityimmunoglobulin TNR6_HUMAN P25445 Tumor necrosis factor MAJOR ISOFORMepsilon FC receptor ISOFORM 5 receptor superfamily member 6 precursorFTCD_HUMAN O95954 Formimidoyltransferase- TPA_HUMAN P00750 Tissue-typeplasminogen ISOFORM E cyclodeaminase ISOFORM activator precursor SHORTFUT8_HUMAN Q9BYC5 Alpha-(1,6)-fucosyltransferase TPO_HUMAN P40225Thrombopoietin precursor ISOFORM 2 MAJOR ISOFORM FXM1_HUMAN Q08050Forkhead box protein M1 TR12_HUMAN Q99831 Tumor necrosis factor ISOFORM2 ISOFORM 12 receptor superfamily member 12 precursor G72_HUMAN P59103Protein G72 TR12_HUMAN Q99831 Tumor necrosis factor MAJOR ISOFORMISOFORM 3 receptor superfamily member 12 precursor G72_HUMAN P59103Protein G72 TR12_HUMAN Q99831 Tumor necrosis factor ISOFORM 2 ISOFORM 4receptor superfamily member 12 precursor G8_HUMAN Q9UBA6 G8 proteinTR12_HUMAN Q99831 Tumor necrosis factor MAJOR ISOFORM ISOFORM 5 receptorsuperfamily member 12 precursor GBR1_HUMAN Q9UBS5 Gamma-aminobutyricacid type B TR12_HUMAN Q99831 Tumor necrosis factor ISOFORM 1E receptorsubunit 1 precursor ISOFORM 6 receptor superfamily member 12 precursorGCFC_HUMAN Q9Y5B6 GC-rich sequence DNA-binding TR12_HUMAN Q99831 Tumornecrosis factor ISOFORM D factor homolog ISOFORM 7 receptor superfamilymember 12 precursor GCP2_HUMAN Q9BSJ2 Gamma-tubulin complex U713_HUMANQ9BYM8 Ubiquitin conjugating ISOFORM 2 component 2 ISOFORM 2 enzyme 7interacting protein 3 GDNR_HUMAN P56159 GDNF receptor alpha precursorU713_HUMAN Q9BYM8 Ubiquitin conjugating ISOFORM 2 ISOFORM 4 enzyme 7interacting protein 3 GLI2_HUMAN P10070 Zinc finger protein GLI2USH3_HUMAN P58418 Usher syndrome type 3 MAJOR ISOFORM ISOFORM B proteinGLI2_HUMAN P10070 Zinc finger protein GLI2 WS14_HUMAN Q9NP71Williams-Beuren ISOFORM BETA ISOFORM 5 syndrome chromosome region 14protein GLI2_HUMAN P10070 Zinc finger protein GLI2 XE7_HUMAN Q02040B-lymphocyte antigen ISOFORM DELTA ISOFORM precursor SHORT GLI2_HUMANP10070 Zinc finger protein GLI2 Z236_HUMAN Q9UL36 Zinc finger protein236 ISOFORM ISOFORM A GAMMA GLSK_HUMAN O94925 Glutaminase kidney isoformZAN_HUMAN Q9BXN9 Zonadhesin precursor ISOFORM GAC mitochondrialprecursor ISOFORM 1 GPT_HUMAN Q9H3H5 UDP-N-acetylglucosamine-- ZAN_HUMANQ9BXN9 Zonadhesin precursor MAJOR ISOFORM dolichyl-phosphate N- ISOFORM2 GRB2_HUMAN P29354 Growth factor receptor-bound ZAN_HUMAN Q9BXN9Zonadhesin precursor MAJOR ISOFORM protein 2 ISOFORM 4 HAIR_HUMAN O43593Hairless protein ZAN_HUMAN Q9BXN9 Zonadhesin precursor MAJOR ISOFORMISOFORM 5 HAIR_HUMAN O43593 Hairless protein ISOFORM SHORT

1. A method for systematically characterizing putative protein isoformsas apparent targets of nonsense-mediated decay (NMD), the methodcomprising the following steps, each of which is performed with asuitably programmed computer: identifying a dataset of target putativeprotein isoform sequences from a curated protein sequence database forcharacterization; identifying from an mRNA dataset corresponding mRNAsequences representing transcripts encoding the protein isoforms,wherein each of the transcripts comprises a termination codon and afinal intron; determining corresponding gene intron-exon structures bymapping the mRNA sequences to corresponding genomic sequences;determining if the transcripts are apparent targets of NMD by comparingthe position of the termination codon to the position of the finalintron, wherein if the termination codon of the transcript is more than50 nucleotides upstream of the final intron position, then thetranscript is an apparent target of NMD; and outputting from thecomputer a list of said isoforms that are apparent targets of NMD,wherein the dataset of target putative protein isoform sequencescomprises a subset of the isoforms of Table 1: TABLE 1 Protein isoformscharacterized as targets of NMD, ISOFORM NAME ACCESSION PROTEINDESCRIPTION 3BP2_HUMANISO P78314 SH3 domain-binding protein 2 FORM SHORT5H4_HUMANMAJOR Q13639 5-hydroxytryptamine 4 ISOFORM receptorA1A1_HUMANISO P05023 Sodium/potassium-transporting FORM SHORT ATPasealpha-1 chain precursor ABCD_HUMANISO Q9NSE7 Putative ATP-bindingcassette FORM 2 transporter C13 ABCD_HUMANISO Q9NSE7 PutativeATP-binding cassette FORM 3 transporter C13 AD11_HUMANISO O75078 ADAM 11precursor FORM SHORT AD12_HUMANMAJOR O43184 ADAM 12 precursor ISOFORMAD22_HUMANISO Q9POK1 ADAM 22 precursor FORM 2 AKP1_HUMANISO Q92667 Akinase anchor protein 1 FORM 2 mitochondrial precursor ANPB_HUMANISOP20594 Atrial natriuretic peptide FORM SHORT receptor B precursorAS13_HUMANMAJOR Q8WXK3 Ankyrin repeat and SOCS box ISOFORM containingprotein 13 AS13_HUMANISO Q8WXK3 Ankyrin repeat and SOCS box FORM 2containing protein 13 ATF3_HUMANISO P18847 Cyclic-AMP-dependent FORM 2transcription factor ATF-3 ATR_HUMANMAJOR Q9H6X2 Anthrax toxin receptorISOFORM precursor B3G7_HUMANISO Q9NY97 Beta-1 3-galactosyltransferaseFORM 2 7 BC12_HUMANISO Q9HB09 Bcl-2 related proline-rich FORM 2 proteinBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-4 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-5 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-6 precursorC343_HUMANISO Q9HB55 Cytochrome P450 3A43 FORM 4 CA34_HUMANMAJOR Q01955Collagen alpha 3 ISOFORM CA34_HUMANISO Q01955 Collagen alpha 3 FORM 3CAL0_HUMANISO P01258 Calcitonin precursor [Contains: FORM 2 Calcitonin;Katacalcin CANA_HUMANISO Q9HC93 Calpain 10 FORM B CANA_HUMANISO Q9HC93Calpain 10 FORM D CANA_HUMANISO Q9HC93 Calpain 10 FORM E CANA_HUMANISOQ9HC93 Calpain 10 FORM F CBS_HUMANMAJOR P35520 Cystathioninebeta-synthase ISOFORM CD38_HUMANISO P28907 ADP-ribosyl cyclase 1 FORM 2CD47_HUMANISO Q08722 Leukocyte surface antigen FORM OA3-305 CD47precursor CFLA_HUMANISO O43618 CASP8 and FADD-like FORM 9 apoptosisregulator precursor CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 3CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 4 CIQ2_HUMANISO O43526Potassium voltage-gated FORM 3 channel subfamily KQT member 2CIW4_HUMANISO Q9NYG8 Potassium channel subfamily FORM 2 K member 4CLK1_HUMANISO P49759 Protein kinase CLK1 FORM SHORT CLK2_HUMANISO P49760Protein kinase CLK2 FORM SHORT CLK3_HUMANISO P49761 Protein kinase CLK3FORM 2 CML1_HUMANMAJOR Q99788 Chemokine receptor-like 1 ISOFORMCOG4_HUMANISO Q9H9E3 Conserved oilgomeric Golgi FORM 2 complex component4 COLQ_HUMANISO Q9UP88 Acetylcholinesterase FORM VII collagenic tailpeptide precursor CPXM_HUMANISO Q96SM3 Potential carboxypeptidase X FORM2 precursor CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 4CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 5 CT24_HUMANISOQ9BUV8 Protein C20orf24 FORM 4 CTGF_HUMANMAJOR P29279 Connective tissuegrowth ISOFORM factor precursor CU07_HUMANISO P57077 Protein C21orf7FORM B CU07_HUMANISO P57077 Protein C21orf7 FORM C CU18_HUMANISO Q9NVD3Protein C21orf18 FORM B CU63_HUMANISO P58658 Protein C21orf63 precursorFORM B CU80_HUMANISO Q9Y2G5 Protein C21orf80 FORM B CYB5_HUMANISO P00167Cytochrome b5 FORM 2 DACA_HUMANISO Q9BYJ9 Dermatomyositis associatedFORM 2 with cancer putative autoantigen-1 DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM BETA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM DELTA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM GAMMA kDa subunit DJB2_HUMANMAJOR P25686DnaJ homolog subfamily B ISOFORM member 2 DJB2_HUMANISO P25686 DnaJhomolog subfamily B FORM 3 member 2 DONS_HUMANISO Q9NYP3 Downstream ofson gene FORM 2 protein DONS_HUMANIS Q9NYP3 Downstream of son gene OFORM3 protein DPP3_HUMANISO Q9NY33 Dipeptidyl-peptidase III FORM 2DSCA_HUMANISO O60469 Down syndrome cell adhesion FORM SHORT moleculeprecursor DTNB_HUMANISO O60941 Dystrobrevin beta FORM 3 EPA3_HUMANMAJORP29320 Ephrin type-A receptor 3 ISOFORM precursor ERAL_HUMANISO O75616GTP-binding protein era FORM HERA-B homolog ESR2_HUMANISO Q9UHD3Estrogen receptor beta FORM 3 F263_HUMANMAJOR Q16875 6-phosphofructo-2-ISOFORM kinase/fructose-2 6- biphosphatase 3 FAFY_HUMANISO O00507Probable ubiquitin carboxyl- FORM SHORT terminal hydrolase FAF-YFCAR_HUMANISO P24071 Immunoglobulin alpha Fc FORM B-DELTA- receptorprecursor S2 FCE2_HUMANMAJOR P06734 Low affinity immunoglobulin ISOFORMepsilon FC receptor FTCD_HUMANISO O95954 Formimidoyltransferase- FORM Ecyclodeaminase FUT8_HUMANISO Q9BYC5 Alpha-(1,6)-fucosyltransferase FORM2 FXM1_HUMANISO Q08050 Forkhead box protein M1 FORM 2 G72_HUMANMAJORP59103 Protein G72 ISOFORM G72_HUMANISO P59103 Protein G72 FORM 2G8_HUMANMAJOR Q9UBA6 G8 protein ISOFORM GBR1_HUMANISO Q9UBS5Gamma-aminobutyric acid FORM 1E type B receptor subunit 1 precursorGCFC_HUMANISO Q9Y5B6 GC-rich sequence DNA- FORM D binding factor homologGCP2_HUMANISO Q9BSJ2 Gamma-tubulin complex FORM 2 component 2GDNR_HUMANISO P56159 GDNF receptor alpha FORM 2 precursorGLI2_HUMANMAJOR P10070 Zinc finger protein GLI2 ISOFORM GLI2_HUMANISOP10070 Zinc finger protein GLI2 FORM BETA GLI2_HUMANISO P10070 Zincfinger protein GLI2 FORM DELTA GLI2_HUMANISO P10070 Zinc finger proteinGLI2 FORM GAMMA GLSK_HUMANISO O94925 Glutaminase kidney isoform FORM GACmitochondrial precursor GPT_HUMANMAJOR Q9H3H5 UDP-N-acetylglucosamine--ISOFORM dolichyl-phosphate N- GRB2_HUMANMAJOR P29354 Growth factorreceptor-bound ISOFORM protein 2 HAIR_HUMANMAJOR O43593 Hairless proteinISOFORM HAIR_HUMANISO O43593 Hairless protein FORM SHORT HFE_HUMANMAJORQ9HC68 Hereditary ISOFORM hemochromatosis protein precursorI17S_HUMANISO Q9NRM6 Interleukin-17B receptor FORM 2 precursorICE2_HUMANISO P42575 Caspase-2 precursor FORM ICH-1S ICE8_HUMANISOQ9C0K4 Caspase-8 precursor FORM 7 ICEA_HUMANISO Q92851 Caspase-10precursor FORM B ICEA_HUMANISO Q92851 Caspase-10 precursor FORM CILF1_HUMANISO Q01167 Interleukin enhancer- FORM 2 binding factor 1IRF7_HUMANISO Q92985 Interferon regulatory factor 7 FORM C IRL1_HUMANISOQ01638 Interleukin 1 receptor-like FORM C 1 precursor ITP1_HUMANMAJORO14713 Integrin beta-1 binding ISOFORM protein 1 ITP1_HUMANISO O14713Integrin beta-1 binding FORM 2 protein 1 KC11_HUMANIS Q9HCP0 Caseinkinase I gamma 1 OFORM 1S isoform KLK2_HUMANIS P20151 Glandularkallikrein 2 OFORM 3 precursor KLKF_HUMANISO Q9H2R5 Kallikrein 15precursor FORM 2 LEF1_HUMANISO Q9UJU2 Lymphoid enhancer FORM B bindingfactor 1 LFA3_HUMANISO P19256 Lymphocyte function- FORM SHORT associatedantigen 3 precursor LIK1_HUMANISO P53667 LIM domain kinase 1 FORM 3LSHR_HUMANMAJOR P22888 Lutropin- ISOFORM choriogonadotropic hormonereceptor precursor LYST_HUMANMAJOR Q99698 Lysosomal trafficking ISOFORMregulator M2A2_HUMANISO P49641 Alpha-mannosidase IIx FORM SHORTMADI_HUMANISO O95405 Mothers against FORM 2 decapentaplegic homologinteracting protein MAP2_HUMAN P11137 Microtubule-associated MAJORprotein 2 ISOFORM MAP2_HUMANI P11137 Microtubule-associated SOFORM MAP2Cprotein 2 MAP4_HUMANI P27816 Microtubule-associated SOFORM 2 protein 4MAX_HUMANISO P25912 Max protein FORM 3 MK11_HUMANISO Q15759Mitogen-activated protein FORM BETA-2 kinase 11 MLH3_HUMANISO Q9UHC1 DNAmismatch repair FORM 2 protein Mlh3 MRP3_HUMANISO O15438 Canalicularmultispecific FORM 3A organic anion transporter 2 MRP3_HUMANISO O15438Canalicular multispecific FORM 3B organic anion transporter 2MSRE_HUMANISO P21757 Macrophage scavenger FORM II receptor types I andII MTF2_HUMANMAJOR Q9Y483 Metal-response element- ISOFORM bindingtranscription factor 2 NK31_HUMANMAJOR Q99801 Homeobox protein Nkx-31ISOFORM NXF5_HUMANMAJOR Q9H1B4 Nuclear RNA export factor 5 ISOFORMNXFS_HUMANISO Q9H1B4 Nuclear RNA export factor 5 FORM B NXFS_HUMANISOQ9H1B4 Nuclear RNA export factor 5 FORM C NXF5_HUMANISO Q9H1B4 NuclearRNA export factor 5 FORM D NXFS_HUMANISO Q9H1B4 Nuclear RNA exportfactor 5 FORM E PHMX_HUMANISO Q96QS1 Phemx protein FORM 4 PHMX_HUMANISOQ96QS1 Phemx protein FORM 5 PML_HUMANISO P29590 Probable transcriptionFORM PML-3B factor PML PPE1_HUMANISO O14829 Serine/threonine proteinFORM 2 phosphatase with EF- hands-1 PPT2_HUMANISO Q9UMR5Palmitoyl-protein FORM 2 thioesterase 2 precursor PRD7_HUMANMAJOR Q9NQW5PR-domain zinc finger ISOFORM protein 7 PSA7_HUMANISO O14818 Proteasomesubunit alpha FORM 4 type 7 PSD4_HUMANISO P55036 26S proteasome non-FORM RPN10E ATPase regulatory subunit 4 PSN1_HUMANISO P49768 Presenilin1 FORM I-374 PTPD_HUMANMAJOR P23468 Protein-tyrosine ISOFORM phosphatasedelta precursor R51D_HUMANISO O94908 DNA repair protein FORM 2 RAD51homolog 4 RBMS_HUMAN Q93062 RNA-binding protein with MAJOR multiplesplicing ISOFORM RED1_HUMANMAJOR P78563 Double-stranded RNA- ISOFORMspecific editase 1 RHD_HUMANMAJOR Q9UQ21 Blood group Rh ISOFORMRHD_HUMANISO Q9UQ21 Blood group Rh FORM 3 RYK_HUMANMAJOR P34925Tyrosine-protein kinase ISOFORM RYK precursor RYK_HUMANIS P34925Tyrosine-protein kinase OFORM 2 RYK precursor SCA1_ HUMANISO O15126Secretory carrier-associated FORM 2 membrane protein 1 SEN7_HUMANMAJORQ9BQF6 Sentrin-specific protease 7 ISOFORM SFR5_HUMANISO Q13243 Splicingfactor FORM SRP40-2 arginine/serine-rich 5 SHX2_HUMANMAJOR O60902 Shortstature homeobox ISOFORM protein 2 SNB2 HUMANISO Q13425Beta-2-syntrophin FORM 2 SNXD_HUMANISO Q9Y5W8 Sorting nexin 13 FORM 2SON_HUMANISO Q9UPY0 SON protein FORM C SON_HUMANISO Q9UPY0 SON proteinFORM E SUR5_HUMANISO Q15528 Surfeit locus protein 5 FORM SURF5AT10B_HUMANMAJOR O14763 Tumor necrosis factor ISOFORM receptorsuperfamily member 10B precursor T10B HUMANISO O14763 Tumor necrosisfactor FORM SHORT receptor superfamily member 10B precursorTM31_HUMANISO Q9BZY9 Tripartite motif protein 31 FORM BETA TNR6_HUMANISOP25445 Tumor necrosis factor FORM 2 receptor superfamily member 6precursor TNR6_HUMANISO P25445 Tumor necrosis factor FORM 3 receptorsuperfamily member 6 precursor TNR6_HUMANISO P25445 Tumor necrosisfactor FORM 4 receptor superfamily member 6 precursor TNR6_HUMANISOP25445 Tumor necrosis factor FORM 5 receptor superfamily member 6precursor TPA_HUMANISO P00750 Tissue-type plasminogen FORM SHORTactivator precursor TPO HUMANMAJOR P40225 Thrombopoietin precursorISOFORM TR12_HUMANISO Q99831 Tumor necrosis factor FORM 12 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 3 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 4 receptor superfamily member 12precursor TR12_HUMANISO Q99831 Tumor necrosis factor FORM 5 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 6 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 7 receptor superfamily member 12precursor U7I3_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 2 enzyme 7interacting protein 3 U713_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 4enzyme 7 interacting protein 3 USH3_HUMANISO P58418 Usher syndrome type3 FORM B protein WS14_HUMANISO Q9NP71 Williams-Beuren syndrome FORM 5chromosome region 14 protein XE7_HUMANISO Q02040 B-lymphocyte antigenFORM SHORT precursor Z236_HUMANISO Q9UL36 Zinc finger protein 236 FORM AZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM 1 ZAN_HUMANISO Q9BXN9Zonadhesin precursor FORM 2 ZAN_HUMANISO Q9BXN9 Zonadhesin precursorFORM 4 ZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM
 5.


2. The method of claim 1 wherein the dataset of target putative proteinisoform sequences comprises isoforms of Calpain-10, CLK1, CLK2, CLK3,and LARD/TNFRSF12.
 3. A method for systematically characterizingputative protein isoforms as apparent targets of nonsense-mediated decay(NMD), the method comprising the following steps, each of which isperformed with a suitably programmed computer: identifying a dataset oftarget putative protein isoform sequences from a curated proteinsequence database for characterization; identifying corresponding geneintron-exon structures by mapping corresponding mRNA sequences of anmRNA sequence dataset to genomic sequences of a genomic DNA sequencedataset; screening the isoform dataset for a subset of isoforms encodedby transcripts comprising alternate splices which introduce a stop codonmore than 50 nucleotides upstream of the final exon-exon splicejunction; and classifying the subset isoforms as being encoded bytranscripts comprising premature stop codons and as targets of NMD; andoutputting from the computer a list of said isoforms that are apparenttargets of NMD, wherein the dataset of target putative protein isoformsequences comprises a subset of the isoforms of Table 1: TABLE 1 Proteinisoforms characterized as targets of NMD, ISOFORM NAME ACCESSION PROTEINDESCRIPTION 3BP2_HUMANISO P78314 SH3 domain-binding protein 2 FORM SHORT5H4_HUMANMAJOR Q13639 5-hydroxytryptamine 4 ISOFORM receptorA1A1_HUMANISO P05023 Sodium/potassium-transporting FORM SHORT ATPasealpha-1 chain precursor ABCD_HUMANISO Q9NSE7 Putative ATP-bindingcassette FORM 2 transporter C13 ABCD_HUMANISO Q9NSE7 PutativeATP-binding cassette FORM 3 transporter C13 AD11_HUMANISO O75078 ADAM 11precursor FORM SHORT AD12_HUMANMAJOR O43184 ADAM 12 precursor ISOFORMAD22_HUMANISO Q9POK1 ADAM 22 precursor FORM 2 AKP1_HUMANISO Q92667 Akinase anchor protein 1 FORM 2 mitochondrial precursor ANPB_HUMANISOP20594 Atrial natriuretic peptide FORM SHORT receptor B precursorAS13_HUMANMAJOR Q8WXK3 Ankyrin repeat and SOCS box ISOFORM containingprotein 13 AS13_HUMANISO Q8WXK3 Ankyrin repeat and SOCS box FORM 2containing protein 13 ATF3_HUMANISO P18847 Cyclic-AMP-dependent FORM 2transcription factor ATF-3 ATR_HUMANMAJOR Q9H6X2 Anthrax toxin receptorISOFORM precursor B3G7_HUMANISO Q9NY97 Beta-1 3-galactosyltransferaseFORM 2 7 BC12_HUMANISO Q9HB09 Bcl-2 related proline-rich FORM 2 proteinBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-4 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-5 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-6 precursorC343_HUMANISO Q9HB55 Cytochrome P450 3A43 FORM 4 CA34_HUMANMAJOR Q01955Collagen alpha 3 ISOFORM CA34_HUMANISO Q01955 Collagen alpha 3 FORM 3CAL0_HUMANISO P01258 Calcitonin precursor [Contains: FORM 2 Calcitonin;Katacalcin CANA_HUMANISO Q9HC93 Calpain 10 FORM B CANA_HUMANISO Q9HC93Calpain 10 FORM D CANA_HUMANISO Q9HC93 Calpain 10 FORM E CANA_HUMANISOQ9HC93 Calpain 10 FORM F CBS_HUMANMAJOR P35520 Cystathioninebeta-synthase ISOFORM CD38_HUMANISO P28907 ADP-ribosyl cyclase 1 FORM 2CD47_HUMANISO Q08722 Leukocyte surface antigen FORM OA3-305 CD47precursor CFLA_HUMANISO O43618 CASP8 and FADD-like FORM 9 apoptosisregulator precursor CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 3CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 4 CIQ2_HUMANISO O43526Potassium voltage-gated FORM 3 channel subfamily KQT member 2CIW4_HUMANISO Q9NYG8 Potassium channel subfamily FORM 2 K member 4CLK1_HUMANISO P49759 Protein kinase CLK1 FORM SHORT CLK2_HUMANISO P49760Protein kinase CLK2 FORM SHORT CLK3_HUMANISO P49761 Protein kinase CLK3FORM 2 CML1_HUMANMAJOR Q99788 Chemokine receptor-like 1 ISOFORMCOG4_HUMANISO Q9H9E3 Conserved oilgomeric Golgi FORM 2 complex component4 COLQ_HUMANISO Q9UP88 Acetylcholinesterase FORM VII collagenic tailpeptide precursor CPXM_HUMANISO Q96SM3 Potential carboxypeptidase X FORM2 precursor CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 4CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 5 CT24_HUMANISOQ9BUV8 Protein C20orf24 FORM 4 CTGF_HUMANMAJOR P29279 Connective tissuegrowth ISOFORM factor precursor CU07_HUMANISO P57077 Protein C21orf7FORM B CU07_HUMANISO P57077 Protein C21orf7 FORM C CU18_HUMANISO Q9NVD3Protein C21orf18 FORM B CU63_HUMANISO P58658 Protein C21orf63 precursorFORM B CU80_HUMANISO Q9Y2G5 Protein C21orf80 FORM B CYB5_HUMANISO P00167Cytochrome b5 FORM 2 DACA_HUMANISO Q9BYJ9 Dermatomyositis associatedFORM 2 with cancer putative autoantigen-1 DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM BETA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM DELTA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM GAMMA kDa subunit DJB2_HUMANMAJOR P25686DnaJ homolog subfamily B ISOFORM member 2 DJB2_HUMANISO P25686 DnaJhomolog subfamily B FORM 3 member 2 DONS_HUMANISO Q9NYP3 Downstream ofson gene FORM 2 protein DONS_HUMANIS Q9NYP3 Downstream of son gene OFORM3 protein DPP3_HUMANISO Q9NY33 Dipeptidyl-peptidase III FORM 2DSCA_HUMANISO O60469 Down syndrome cell adhesion FORM SHORT moleculeprecursor DTNB_HUMANISO O60941 Dystrobrevin beta FORM 3 EPA3_HUMANMAJORP29320 Ephrin type-A receptor 3 ISOFORM precursor ERAL_HUMANISO O75616GTP-binding protein era FORM HERA-B homolog ESR2_HUMANISO Q9UHD3Estrogen receptor beta FORM 3 F263_HUMANMAJOR Q16875 6-phosphofructo-2-ISOFORM kinase/fructose-2 6- biphosphatase 3 FAFY_HUMANISO O00507Probable ubiquitin carboxyl- FORM SHORT terminal hydrolase FAF-YFCAR_HUMANISO P24071 Immunoglobulin alpha Fc FORM B-DELTA- receptorprecursor S2 FCE2_HUMANMAJOR P06734 Low affinity immunoglobulin ISOFORMepsilon FC receptor FTCD_HUMANISO O95954 Formimidoyltransferase- FORM Ecyclodeaminase FUT8_HUMANISO Q9BYC5 Alpha-(1,6)-fucosyltransferase FORM2 FXM1_HUMANISO Q08050 Forkhead box protein M1 FORM 2 G72_HUMANMAJORP59103 Protein G72 ISOFORM G72_HUMANISO P59103 Protein G72 FORM 2G8_HUMANMAJOR Q9UBA6 G8 protein ISOFORM GBR1_HUMANISO Q9UBS5Gamma-aminobutyric acid FORM 1E type B receptor subunit 1 precursorGCFC_HUMANISO Q9Y5B6 GC-rich sequence DNA- FORM D binding factor homologGCP2_HUMANISO Q9BSJ2 Gamma-tubulin complex FORM 2 component 2GDNR_HUMANISO P56159 GDNF receptor alpha FORM 2 precursorGLI2_HUMANMAJOR P10070 Zinc finger protein GLI2 ISOFORM GLI2_HUMANISOP10070 Zinc finger protein GLI2 FORM BETA GLI2_HUMANISO P10070 Zincfinger protein GLI2 FORM DELTA GLI2_HUMANISO P10070 Zinc finger proteinGLI2 FORM GAMMA GLSK_HUMANISO O94925 Glutaminase kidney isoform FORM GACmitochondrial precursor GPT_HUMANMAJOR Q9H3H5 UDP-N-acetylglucosamine--ISOFORM dolichyl-phosphate N- GRB2_HUMANMAJOR P29354 Growth factorreceptor-bound ISOFORM protein 2 HAIR_HUMANMAJOR O43593 Hairless proteinISOFORM HAIR_HUMANISO O43593 Hairless protein FORM SHORT HFE_HUMANMAJORQ9HC68 Hereditary ISOFORM hemochromatosis protein precursorI17S_HUMANISO Q9NRM6 Interleukin-17B receptor FORM 2 precursorICE2_HUMANISO P42575 Caspase-2 precursor FORM ICH-1S ICE8_HUMANISOQ9C0K4 Caspase-8 precursor FORM 7 ICEA_HUMANISO Q92851 Caspase-10precursor FORM B ICEA_HUMANISO Q92851 Caspase-10 precursor FORM CILF1_HUMANISO Q01167 Interleukin enhancer- FORM 2 binding factor 1IRF7_HUMANISO Q92985 Interferon regulatory factor 7 FORM C IRL1_HUMANISOQ01638 Interleukin 1 receptor-like FORM C 1 precursor ITP1_HUMANMAJORO14713 Integrin beta-1 binding ISOFORM protein 1 ITP1_HUMANISO O14713Integrin beta-1 binding FORM 2 protein 1 KC11_HUMANIS Q9HCP0 Caseinkinase I gamma 1 OFORM 1S isoform KLK2_HUMANIS P20151 Glandularkallikrein 2 OFORM 3 precursor KLKF_HUMANISO Q9H2R5 Kallikrein 15precursor FORM 2 LEF1_HUMANISO Q9UJU2 Lymphoid enhancer FORM B bindingfactor 1 LFA3_HUMANISO P19256 Lymphocyte function- FORM SHORT associatedantigen 3 precursor LIK1_HUMANISO P53667 LIM domain kinase 1 FORM 3LSHR_HUMANMAJOR P22888 Lutropin- ISOFORM choriogonadotropic hormonereceptor precursor LYST_HUMANMAJOR Q99698 Lysosomal trafficking ISOFORMregulator M2A2_HUMANISO P49641 Alpha-mannosidase IIx FORM SHORTMADI_HUMANISO O95405 Mothers against FORM 2 decapentaplegic homologinteracting protein MAP2_HUMAN P11137 Microtubule-associated MAJORprotein 2 ISOFORM MAP2_HUMANI P11137 Microtubule-associated SOFORM MAP2Cprotein 2 MAP4_HUMANI P27816 Microtubule-associated SOFORM 2 protein 4MAX_HUMANISO P25912 Max protein FORM 3 MK11_HUMANISO Q15759Mitogen-activated protein FORM BETA-2 kinase 11 MLH3_HUMANISO Q9UHC1 DNAmismatch repair FORM 2 protein Mlh3 MRP3_HUMANISO O15438 Canalicularmultispecific FORM 3A organic anion transporter 2 MRP3_HUMANISO O15438Canalicular multispecific FORM 3B organic anion transporter 2MSRE_HUMANISO P21757 Macrophage scavenger FORM II receptor types I andII MTF2_HUMANMAJOR Q9Y483 Metal-response element- ISOFORM bindingtranscription factor 2 NK31_HUMANMAJOR Q99801 Homeobox protein Nkx-31ISOFORM NXF5_HUMANMAJOR Q9H1B4 Nuclear RNA export factor 5 ISOFORMNXFS_HUMANISO Q9H1B4 Nuclear RNA export factor 5 FORM B NXFS_HUMANISOQ9H1B4 Nuclear RNA export factor 5 FORM C NXF5_HUMANISO Q9H1B4 NuclearRNA export factor 5 FORM D NXFS_HUMANISO Q9H1B4 Nuclear RNA exportfactor 5 FORM E PHMX_HUMANISO Q96QS1 Phemx protein FORM 4 PHMX_HUMANISOQ96QS1 Phemx protein FORM 5 PML_HUMANISO P29590 Probable transcriptionFORM PML-3B factor PML PPE1_HUMANISO O14829 Serine/threonine proteinFORM 2 phosphatase with EF- hands-1 PPT2_HUMANISO Q9UMR5Palmitoyl-protein FORM 2 thioesterase 2 precursor PRD7_HUMANMAJOR Q9NQW5PR-domain zinc finger ISOFORM protein 7 PSA7_HUMANISO O14818 Proteasomesubunit alpha FORM 4 type 7 PSD4_HUMANISO P55036 26S proteasome non-FORM RPN10E ATPase regulatory subunit 4 PSN1_HUMANISO P49768 Presenilin1 FORM I-374 PTPD_HUMANMAJOR P23468 Protein-tyrosine ISOFORM phosphatasedelta precursor R51D_HUMANISO O94908 DNA repair protein FORM 2 RAD51homolog 4 RBMS_HUMAN Q93062 RNA-binding protein with MAJOR multiplesplicing ISOFORM RED1_HUMANMAJOR P78563 Double-stranded RNA- ISOFORMspecific editase 1 RHD_HUMANMAJOR Q9UQ21 Blood group Rh ISOFORMRHD_HUMANISO Q9UQ21 Blood group Rh FORM 3 RYK_HUMANMAJOR P34925Tyrosine-protein kinase ISOFORM RYK precursor RYK_HUMANIS P34925Tyrosine-protein kinase OFORM 2 RYK precursor SCA1_ HUMANISO O15126Secretory carrier-associated FORM 2 membrane protein 1 SEN7_HUMANMAJORQ9BQF6 Sentrin-specific protease 7 ISOFORM SFR5_HUMANISO Q13243 Splicingfactor FORM SRP40-2 arginine/serine-rich 5 SHX2_HUMANMAJOR O60902 Shortstature homeobox ISOFORM protein 2 SNB2 HUMANISO Q13425Beta-2-syntrophin FORM 2 SNXD_HUMANISO Q9Y5W8 Sorting nexin 13 FORM 2SON_HUMANISO Q9UPY0 SON protein FORM C SON_HUMANISO Q9UPY0 SON proteinFORM E SUR5_HUMANISO Q15528 Surfeit locus protein 5 FORM SURF5AT10B_HUMANMAJOR O14763 Tumor necrosis factor ISOFORM receptorsuperfamily member 10B precursor T10B HUMANISO O14763 Tumor necrosisfactor FORM SHORT receptor superfamily member 10B precursorTM31_HUMANISO Q9BZY9 Tripartite motif protein 31 FORM BETA TNR6_HUMANISOP25445 Tumor necrosis factor FORM 2 receptor superfamily member 6precursor TNR6_HUMANISO P25445 Tumor necrosis factor FORM 3 receptorsuperfamily member 6 precursor TNR6_HUMANISO P25445 Tumor necrosisfactor FORM 4 receptor superfamily member 6 precursor TNR6_HUMANISOP25445 Tumor necrosis factor FORM 5 receptor superfamily member 6precursor TPA_HUMANISO P00750 Tissue-type plasminogen FORM SHORTactivator precursor TPO HUMANMAJOR P40225 Thrombopoietin precursorISOFORM TR12_HUMANISO Q99831 Tumor necrosis factor FORM 12 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 3 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 4 receptor superfamily member 12precursor TR12_HUMANISO Q99831 Tumor necrosis factor FORM 5 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 6 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 7 receptor superfamily member 12precursor U7I3_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 2 enzyme 7interacting protein 3 U713_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 4enzyme 7 interacting protein 3 USH3_HUMANISO P58418 Usher syndrome type3 FORM B protein WS14_HUMANISO Q9NP71 Williams-Beuren syndrome FORM 5chromosome region 14 protein XE7_HUMANISO Q02040 B-lymphocyte antigenFORM SHORT precursor Z236_HUMANISO Q9UL36 Zinc finger protein 236 FORM AZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM 1 ZAN_HUMANISO Q9BXN9Zonadhesin precursor FORM 2 ZAN_HUMANISO Q9BXN9 Zonadhesin precursorFORM 4 ZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM
 5.


4. The method of claim 3 wherein the dataset of target putative proteinisoform sequences comprises isoforms of Calpain-10, CLK1, CLK2, CLK3,and LARD/TNFRSF12.
 5. A computer program product, comprising a computerusable medium having a computer readable program code embodied therein,said computer readable program code adapted to be executed to implementa method for systematically characterizing putative protein isoforms asapparent targets of nonsense-mediated decay (NMD) and for generating andoutputting a list of said isoforms, the method comprising the followingsteps: identifying a dataset of target putative protein isoformsequences from a curated protein sequence database for characterization;identifying from an mRNA dataset corresponding mRNA sequencesrepresenting transcripts encoding the protein isoforms, wherein each ofthe transcripts comprises a termination codon and a final intron;determining corresponding gene intron-exon structures by mapping themRNA sequences to corresponding genomic sequences; determining if thetranscripts are apparent targets of NMD by comparing the position of thetermination codon to the position of the final intron, wherein if thetermination codon of the transcript is more than 50 nucleotides upstreamof the final intron position, then the transcript is an apparent targetof NMD; and outputting from the computer a list of said isoforms thatare apparent targets of NMD, wherein the dataset of target putativeprotein isoform sequences comprises a subset of the isoforms of Table 1:TABLE 1 Protein isoforms characterized as targets of NMD, ISOFORM NAMEACCESSION PROTEIN DESCRIPTION 3BP2_HUMANISO P78314 SH3 domain-bindingprotein 2 FORM SHORT 5H4_HUMANMAJOR Q13639 5-hydroxytryptamine 4 ISOFORMreceptor A1A1_HUMANISO P05023 Sodium/potassium-transporting FORM SHORTATPase alpha-1 chain precursor ABCD_HUMANISO Q9NSE7 Putative ATP-bindingcassette FORM 2 transporter C13 ABCD_HUMANISO Q9NSE7 PutativeATP-binding cassette FORM 3 transporter C13 AD11_HUMANISO O75078 ADAM 11precursor FORM SHORT AD12_HUMANMAJOR O43184 ADAM 12 precursor ISOFORMAD22_HUMANISO Q9POK1 ADAM 22 precursor FORM 2 AKP1_HUMANISO Q92667 Akinase anchor protein 1 FORM 2 mitochondrial precursor ANPB_HUMANISOP20594 Atrial natriuretic peptide FORM SHORT receptor B precursorAS13_HUMANMAJOR Q8WXK3 Ankyrin repeat and SOCS box ISOFORM containingprotein 13 AS13_HUMANISO Q8WXK3 Ankyrin repeat and SOCS box FORM 2containing protein 13 ATF3_HUMANISO P18847 Cyclic-AMP-dependent FORM 2transcription factor ATF-3 ATR_HUMANMAJOR Q9H6X2 Anthrax toxin receptorISOFORM precursor B3G7_HUMANISO Q9NY97 Beta-1 3-galactosyltransferaseFORM 2 7 BC12_HUMANISO Q9HB09 Bcl-2 related proline-rich FORM 2 proteinBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-4 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-5 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-6 precursorC343_HUMANISO Q9HB55 Cytochrome P450 3A43 FORM 4 CA34_HUMANMAJOR Q01955Collagen alpha 3 ISOFORM CA34_HUMANISO Q01955 Collagen alpha 3 FORM 3CAL0_HUMANISO P01258 Calcitonin precursor [Contains: FORM 2 Calcitonin;Katacalcin CANA_HUMANISO Q9HC93 Calpain 10 FORM B CANA_HUMANISO Q9HC93Calpain 10 FORM D CANA_HUMANISO Q9HC93 Calpain 10 FORM E CANA_HUMANISOQ9HC93 Calpain 10 FORM F CBS_HUMANMAJOR P35520 Cystathioninebeta-synthase ISOFORM CD38_HUMANISO P28907 ADP-ribosyl cyclase 1 FORM 2CD47_HUMANISO Q08722 Leukocyte surface antigen FORM OA3-305 CD47precursor CFLA_HUMANISO O43618 CASP8 and FADD-like FORM 9 apoptosisregulator precursor CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 3CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 4 CIQ2_HUMANISO O43526Potassium voltage-gated FORM 3 channel subfamily KQT member 2CIW4_HUMANISO Q9NYG8 Potassium channel subfamily FORM 2 K member 4CLK1_HUMANISO P49759 Protein kinase CLK1 FORM SHORT CLK2_HUMANISO P49760Protein kinase CLK2 FORM SHORT CLK3_HUMANISO P49761 Protein kinase CLK3FORM 2 CML1_HUMANMAJOR Q99788 Chemokine receptor-like 1 ISOFORMCOG4_HUMANISO Q9H9E3 Conserved oilgomeric Golgi FORM 2 complex component4 COLQ_HUMANISO Q9UP88 Acetylcholinesterase FORM VII collagenic tailpeptide precursor CPXM_HUMANISO Q96SM3 Potential carboxypeptidase X FORM2 precursor CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 4CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 5 CT24_HUMANISOQ9BUV8 Protein C20orf24 FORM 4 CTGF_HUMANMAJOR P29279 Connective tissuegrowth ISOFORM factor precursor CU07_HUMANISO P57077 Protein C21orf7FORM B CU07_HUMANISO P57077 Protein C21orf7 FORM C CU18_HUMANISO Q9NVD3Protein C21orf18 FORM B CU63_HUMANISO P58658 Protein C21orf63 precursorFORM B CU80_HUMANISO Q9Y2G5 Protein C21orf80 FORM B CYB5_HUMANISO P00167Cytochrome b5 FORM 2 DACA_HUMANISO Q9BYJ9 Dermatomyositis associatedFORM 2 with cancer putative autoantigen-1 DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM BETA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM DELTA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM GAMMA kDa subunit DJB2_HUMANMAJOR P25686DnaJ homolog subfamily B ISOFORM member 2 DJB2_HUMANISO P25686 DnaJhomolog subfamily B FORM 3 member 2 DONS_HUMANISO Q9NYP3 Downstream ofson gene FORM 2 protein DONS_HUMANIS Q9NYP3 Downstream of son gene OFORM3 protein DPP3_HUMANISO Q9NY33 Dipeptidyl-peptidase III FORM 2DSCA_HUMANISO O60469 Down syndrome cell adhesion FORM SHORT moleculeprecursor DTNB_HUMANISO O60941 Dystrobrevin beta FORM 3 EPA3_HUMANMAJORP29320 Ephrin type-A receptor 3 ISOFORM precursor ERAL_HUMANISO O75616GTP-binding protein era FORM HERA-B homolog ESR2_HUMANISO Q9UHD3Estrogen receptor beta FORM 3 F263_HUMANMAJOR Q16875 6-phosphofructo-2-ISOFORM kinase/fructose-2 6- biphosphatase 3 FAFY_HUMANISO O00507Probable ubiquitin carboxyl- FORM SHORT terminal hydrolase FAF-YFCAR_HUMANISO P24071 Immunoglobulin alpha Fc FORM B-DELTA- receptorprecursor S2 FCE2_HUMANMAJOR P06734 Low affinity immunoglobulin ISOFORMepsilon FC receptor FTCD_HUMANISO O95954 Formimidoyltransferase- FORM Ecyclodeaminase FUT8_HUMANISO Q9BYC5 Alpha-(1,6)-fucosyltransferase FORM2 FXM1_HUMANISO Q08050 Forkhead box protein M1 FORM 2 G72_HUMANMAJORP59103 Protein G72 ISOFORM G72_HUMANISO P59103 Protein G72 FORM 2G8_HUMANMAJOR Q9UBA6 G8 protein ISOFORM GBR1_HUMANISO Q9UBS5Gamma-aminobutyric acid FORM 1E type B receptor subunit 1 precursorGCFC_HUMANISO Q9Y5B6 GC-rich sequence DNA- FORM D binding factor homologGCP2_HUMANISO Q9BSJ2 Gamma-tubulin complex FORM 2 component 2GDNR_HUMANISO P56159 GDNF receptor alpha FORM 2 precursorGLI2_HUMANMAJOR P10070 Zinc finger protein GLI2 ISOFORM GLI2_HUMANISOP10070 Zinc finger protein GLI2 FORM BETA GLI2_HUMANISO P10070 Zincfinger protein GLI2 FORM DELTA GLI2_HUMANISO P10070 Zinc finger proteinGLI2 FORM GAMMA GLSK_HUMANISO O94925 Glutaminase kidney isoform FORM GACmitochondrial precursor GPT_HUMANMAJOR Q9H3H5 UDP-N-acetylglucosamine--ISOFORM dolichyl-phosphate N- GRB2_HUMANMAJOR P29354 Growth factorreceptor-bound ISOFORM protein 2 HAIR_HUMANMAJOR O43593 Hairless proteinISOFORM HAIR_HUMANISO O43593 Hairless protein FORM SHORT HFE_HUMANMAJORQ9HC68 Hereditary ISOFORM hemochromatosis protein precursorI17S_HUMANISO Q9NRM6 Interleukin-17B receptor FORM 2 precursorICE2_HUMANISO P42575 Caspase-2 precursor FORM ICH-1S ICE8_HUMANISOQ9C0K4 Caspase-8 precursor FORM 7 ICEA_HUMANISO Q92851 Caspase-10precursor FORM B ICEA_HUMANISO Q92851 Caspase-10 precursor FORM CILF1_HUMANISO Q01167 Interleukin enhancer- FORM 2 binding factor 1IRF7_HUMANISO Q92985 Interferon regulatory factor 7 FORM C IRL1_HUMANISOQ01638 Interleukin 1 receptor-like FORM C 1 precursor ITP1_HUMANMAJORO14713 Integrin beta-1 binding ISOFORM protein 1 ITP1_HUMANISO O14713Integrin beta-1 binding FORM 2 protein 1 KC11_HUMANIS Q9HCP0 Caseinkinase I gamma 1 OFORM 1S isoform KLK2_HUMANIS P20151 Glandularkallikrein 2 OFORM 3 precursor KLKF_HUMANISO Q9H2R5 Kallikrein 15precursor FORM 2 LEF1_HUMANISO Q9UJU2 Lymphoid enhancer FORM B bindingfactor 1 LFA3_HUMANISO P19256 Lymphocyte function- FORM SHORT associatedantigen 3 precursor LIK1_HUMANISO P53667 LIM domain kinase 1 FORM 3LSHR_HUMANMAJOR P22888 Lutropin- ISOFORM choriogonadotropic hormonereceptor precursor LYST_HUMANMAJOR Q99698 Lysosomal trafficking ISOFORMregulator M2A2_HUMANISO P49641 Alpha-mannosidase IIx FORM SHORTMADI_HUMANISO O95405 Mothers against FORM 2 decapentaplegic homologinteracting protein MAP2_HUMAN P11137 Microtubule-associated MAJORprotein 2 ISOFORM MAP2_HUMANI P11137 Microtubule-associated SOFORM MAP2Cprotein 2 MAP4_HUMANI P27816 Microtubule-associated SOFORM 2 protein 4MAX_HUMANISO P25912 Max protein FORM 3 MK11_HUMANISO Q15759Mitogen-activated protein FORM BETA-2 kinase 11 MLH3_HUMANISO Q9UHC1 DNAmismatch repair FORM 2 protein Mlh3 MRP3_HUMANISO O15438 Canalicularmultispecific FORM 3A organic anion transporter 2 MRP3_HUMANISO O15438Canalicular multispecific FORM 3B organic anion transporter 2MSRE_HUMANISO P21757 Macrophage scavenger FORM II receptor types I andII MTF2_HUMANMAJOR Q9Y483 Metal-response element- ISOFORM bindingtranscription factor 2 NK31_HUMANMAJOR Q99801 Homeobox protein Nkx-31ISOFORM NXF5_HUMANMAJOR Q9H1B4 Nuclear RNA export factor 5 ISOFORMNXFS_HUMANISO Q9H1B4 Nuclear RNA export factor 5 FORM B NXFS_HUMANISOQ9H1B4 Nuclear RNA export factor 5 FORM C NXF5_HUMANISO Q9H1B4 NuclearRNA export factor 5 FORM D NXFS_HUMANISO Q9H1B4 Nuclear RNA exportfactor 5 FORM E PHMX_HUMANISO Q96QS1 Phemx protein FORM 4 PHMX_HUMANISOQ96QS1 Phemx protein FORM 5 PML_HUMANISO P29590 Probable transcriptionFORM PML-3B factor PML PPE1_HUMANISO O14829 Serine/threonine proteinFORM 2 phosphatase with EF- hands-1 PPT2_HUMANISO Q9UMR5Palmitoyl-protein FORM 2 thioesterase 2 precursor PRD7_HUMANMAJOR Q9NQW5PR-domain zinc finger ISOFORM protein 7 PSA7_HUMANISO O14818 Proteasomesubunit alpha FORM 4 type 7 PSD4_HUMANISO P55036 26S proteasome non-FORM RPN10E ATPase regulatory subunit 4 PSN1_HUMANISO P49768 Presenilin1 FORM I-374 PTPD_HUMANMAJOR P23468 Protein-tyrosine ISOFORM phosphatasedelta precursor R51D_HUMANISO O94908 DNA repair protein FORM 2 RAD51homolog 4 RBMS_HUMAN Q93062 RNA-binding protein with MAJOR multiplesplicing ISOFORM RED1_HUMANMAJOR P78563 Double-stranded RNA- ISOFORMspecific editase 1 RHD_HUMANMAJOR Q9UQ21 Blood group Rh ISOFORMRHD_HUMANISO Q9UQ21 Blood group Rh FORM 3 RYK_HUMANMAJOR P34925Tyrosine-protein kinase ISOFORM RYK precursor RYK_HUMANIS P34925Tyrosine-protein kinase OFORM 2 RYK precursor SCA1_ HUMANISO O15126Secretory carrier-associated FORM 2 membrane protein 1 SEN7_HUMANMAJORQ9BQF6 Sentrin-specific protease 7 ISOFORM SFR5_HUMANISO Q13243 Splicingfactor FORM SRP40-2 arginine/serine-rich 5 SHX2_HUMANMAJOR O60902 Shortstature homeobox ISOFORM protein 2 SNB2 HUMANISO Q13425Beta-2-syntrophin FORM 2 SNXD_HUMANISO Q9Y5W8 Sorting nexin 13 FORM 2SON_HUMANISO Q9UPY0 SON protein FORM C SON_HUMANISO Q9UPY0 SON proteinFORM E SUR5_HUMANISO Q15528 Surfeit locus protein 5 FORM SURF5AT10B_HUMANMAJOR O14763 Tumor necrosis factor ISOFORM receptorsuperfamily member 10B precursor T10B HUMANISO O14763 Tumor necrosisfactor FORM SHORT receptor superfamily member 10B precursorTM31_HUMANISO Q9BZY9 Tripartite motif protein 31 FORM BETA TNR6_HUMANISOP25445 Tumor necrosis factor FORM 2 receptor superfamily member 6precursor TNR6_HUMANISO P25445 Tumor necrosis factor FORM 3 receptorsuperfamily member 6 precursor TNR6_HUMANISO P25445 Tumor necrosisfactor FORM 4 receptor superfamily member 6 precursor TNR6_HUMANISOP25445 Tumor necrosis factor FORM 5 receptor superfamily member 6precursor TPA_HUMANISO P00750 Tissue-type plasminogen FORM SHORTactivator precursor TPO HUMANMAJOR P40225 Thrombopoietin precursorISOFORM TR12_HUMANISO Q99831 Tumor necrosis factor FORM 12 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 3 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 4 receptor superfamily member 12precursor TR12_HUMANISO Q99831 Tumor necrosis factor FORM 5 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 6 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 7 receptor superfamily member 12precursor U7I3_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 2 enzyme 7interacting protein 3 U713_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 4enzyme 7 interacting protein 3 USH3_HUMANISO P58418 Usher syndrome type3 FORM B protein WS14_HUMANISO Q9NP71 Williams-Beuren syndrome FORM 5chromosome region 14 protein XE7_HUMANISO Q02040 B-lymphocyte antigenFORM SHORT precursor Z236_HUMANISO Q9UL36 Zinc finger protein 236 FORM AZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM 1 ZAN_HUMANISO Q9BXN9Zonadhesin precursor FORM 2 ZAN_HUMANISO Q9BXN9 Zonadhesin precursorFORM 4 ZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM
 5.


6. The product of claim 5 wherein the dataset of target putative proteinisoform sequences comprises isoforms of Calpain-10, CLK1, CLK2, CLK3,and LARD/TNFRSF12.
 7. A computer program product, comprising a computerusable medium having a computer readable program code embodied therein,said computer readable program code adapted to be executed to implementa method for systematically characterizing putative protein isoforms asapparent targets of nonsense-mediated decay (NMD) and for generating andoutputting a list of said isoforms, the method comprising the followingsteps: identifying a dataset of target putative protein isoformsequences from a curated protein sequence database for characterization;identifying corresponding gene intron-exon structures by mappingcorresponding mRNA sequences of an mRNA sequence dataset to genomicsequences of a genomic DNA sequence dataset; screening the isoformdataset for a subset of isoforms encoded by transcripts comprisingalternate splices which introduce a stop codon more than 50 nucleotidesupstream of the final exon-exon splice junction; and classifying thesubset isoforms as being encoded by transcripts comprising prematurestop codons and as targets of NMD; and outputting from the computer alist of said isoforms that are apparent targets of NMD, wherein thedataset of target putative protein isoform sequences comprises a subsetof the isoforms of Table 1: TABLE 1 Protein isoforms characterized astargets of NMD, ISOFORM NAME ACCESSION PROTEIN DESCRIPTION 3BP2_HUMANISOP78314 SH3 domain-binding protein 2 FORM SHORT 5H4_HUMANMAJOR Q136395-hydroxytryptamine 4 ISOFORM receptor A1A1_HUMANISO P05023Sodium/potassium-transporting FORM SHORT ATPase alpha-1 chain precursorABCD_HUMANISO Q9NSE7 Putative ATP-binding cassette FORM 2 transporterC13 ABCD_HUMANISO Q9NSE7 Putative ATP-binding cassette FORM 3transporter C13 AD11_HUMANISO O75078 ADAM 11 precursor FORM SHORTAD12_HUMANMAJOR O43184 ADAM 12 precursor ISOFORM AD22_HUMANISO Q9POK1ADAM 22 precursor FORM 2 AKP1_HUMANISO Q92667 A kinase anchor protein 1FORM 2 mitochondrial precursor ANPB_HUMANISO P20594 Atrial natriureticpeptide FORM SHORT receptor B precursor AS13_HUMANMAJOR Q8WXK3 Ankyrinrepeat and SOCS box ISOFORM containing protein 13 AS13_HUMANISO Q8WXK3Ankyrin repeat and SOCS box FORM 2 containing protein 13 ATF3_HUMANISOP18847 Cyclic-AMP-dependent FORM 2 transcription factor ATF-3ATR_HUMANMAJOR Q9H6X2 Anthrax toxin receptor ISOFORM precursorB3G7_HUMANISO Q9NY97 Beta-1 3-galactosyltransferase FORM 2 7BC12_HUMANISO Q9HB09 Bcl-2 related proline-rich FORM 2 proteinBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-4 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-5 precursorBMP1_HUMANISO P13497 Bone morphogenetic protein 1 FORM BMP1-6 precursorC343_HUMANISO Q9HB55 Cytochrome P450 3A43 FORM 4 CA34_HUMANMAJOR Q01955Collagen alpha 3 ISOFORM CA34_HUMANISO Q01955 Collagen alpha 3 FORM 3CAL0_HUMANISO P01258 Calcitonin precursor [Contains: FORM 2 Calcitonin;Katacalcin CANA_HUMANISO Q9HC93 Calpain 10 FORM B CANA_HUMANISO Q9HC93Calpain 10 FORM D CANA_HUMANISO Q9HC93 Calpain 10 FORM E CANA_HUMANISOQ9HC93 Calpain 10 FORM F CBS_HUMANMAJOR P35520 Cystathioninebeta-synthase ISOFORM CD38_HUMANISO P28907 ADP-ribosyl cyclase 1 FORM 2CD47_HUMANISO Q08722 Leukocyte surface antigen FORM OA3-305 CD47precursor CFLA_HUMANISO O43618 CASP8 and FADD-like FORM 9 apoptosisregulator precursor CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 3CHRD_HUMANISO Q9P0Z5 Chordin precursor FORM 4 CIQ2_HUMANISO O43526Potassium voltage-gated FORM 3 channel subfamily KQT member 2CIW4_HUMANISO Q9NYG8 Potassium channel subfamily FORM 2 K member 4CLK1_HUMANISO P49759 Protein kinase CLK1 FORM SHORT CLK2_HUMANISO P49760Protein kinase CLK2 FORM SHORT CLK3_HUMANISO P49761 Protein kinase CLK3FORM 2 CML1_HUMANMAJOR Q99788 Chemokine receptor-like 1 ISOFORMCOG4_HUMANISO Q9H9E3 Conserved oilgomeric Golgi FORM 2 complex component4 COLQ_HUMANISO Q9UP88 Acetylcholinesterase FORM VII collagenic tailpeptide precursor CPXM_HUMANISO Q96SM3 Potential carboxypeptidase X FORM2 precursor CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 4CRN1_HUMANISO Q9NYD8 Crooked neck-like protein 1 FORM 5 CT24_HUMANISOQ9BUV8 Protein C20orf24 FORM 4 CTGF_HUMANMAJOR P29279 Connective tissuegrowth ISOFORM factor precursor CU07_HUMANISO P57077 Protein C21orf7FORM B CU07_HUMANISO P57077 Protein C21orf7 FORM C CU18_HUMANISO Q9NVD3Protein C21orf18 FORM B CU63_HUMANISO P58658 Protein C21orf63 precursorFORM B CU80_HUMANISO Q9Y2G5 Protein C21orf80 FORM B CYB5_HUMANISO P00167Cytochrome b5 FORM 2 DACA_HUMANISO Q9BYJ9 Dermatomyositis associatedFORM 2 with cancer putative autoantigen-1 DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM BETA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM DELTA kDa subunit DFFB_HUMANISO O76075 DNAfragmentation factor 40 FORM GAMMA kDa subunit DJB2_HUMANMAJOR P25686DnaJ homolog subfamily B ISOFORM member 2 DJB2_HUMANISO P25686 DnaJhomolog subfamily B FORM 3 member 2 DONS_HUMANISO Q9NYP3 Downstream ofson gene FORM 2 protein DONS_HUMANIS Q9NYP3 Downstream of son gene OFORM3 protein DPP3_HUMANISO Q9NY33 Dipeptidyl-peptidase III FORM 2DSCA_HUMANISO O60469 Down syndrome cell adhesion FORM SHORT moleculeprecursor DTNB_HUMANISO O60941 Dystrobrevin beta FORM 3 EPA3_HUMANMAJORP29320 Ephrin type-A receptor 3 ISOFORM precursor ERAL_HUMANISO O75616GTP-binding protein era FORM HERA-B homolog ESR2_HUMANISO Q9UHD3Estrogen receptor beta FORM 3 F263_HUMANMAJOR Q16875 6-phosphofructo-2-ISOFORM kinase/fructose-2 6- biphosphatase 3 FAFY_HUMANISO O00507Probable ubiquitin carboxyl- FORM SHORT terminal hydrolase FAF-YFCAR_HUMANISO P24071 Immunoglobulin alpha Fc FORM B-DELTA- receptorprecursor S2 FCE2_HUMANMAJOR P06734 Low affinity immunoglobulin ISOFORMepsilon FC receptor FTCD_HUMANISO O95954 Formimidoyltransferase- FORM Ecyclodeaminase FUT8_HUMANISO Q9BYC5 Alpha-(1,6)-fucosyltransferase FORM2 FXM1_HUMANISO Q08050 Forkhead box protein M1 FORM 2 G72_HUMANMAJORP59103 Protein G72 ISOFORM G72_HUMANISO P59103 Protein G72 FORM 2G8_HUMANMAJOR Q9UBA6 G8 protein ISOFORM GBR1_HUMANISO Q9UBS5Gamma-aminobutyric acid FORM 1E type B receptor subunit 1 precursorGCFC_HUMANISO Q9Y5B6 GC-rich sequence DNA- FORM D binding factor homologGCP2_HUMANISO Q9BSJ2 Gamma-tubulin complex FORM 2 component 2GDNR_HUMANISO P56159 GDNF receptor alpha FORM 2 precursorGLI2_HUMANMAJOR P10070 Zinc finger protein GLI2 ISOFORM GLI2_HUMANISOP10070 Zinc finger protein GLI2 FORM BETA GLI2_HUMANISO P10070 Zincfinger protein GLI2 FORM DELTA GLI2_HUMANISO P10070 Zinc finger proteinGLI2 FORM GAMMA GLSK_HUMANISO O94925 Glutaminase kidney isoform FORM GACmitochondrial precursor GPT_HUMANMAJOR Q9H3H5 UDP-N-acetylglucosamine--ISOFORM dolichyl-phosphate N- GRB2_HUMANMAJOR P29354 Growth factorreceptor-bound ISOFORM protein 2 HAIR_HUMANMAJOR O43593 Hairless proteinISOFORM HAIR_HUMANISO O43593 Hairless protein FORM SHORT HFE_HUMANMAJORQ9HC68 Hereditary ISOFORM hemochromatosis protein precursorI17S_HUMANISO Q9NRM6 Interleukin-17B receptor FORM 2 precursorICE2_HUMANISO P42575 Caspase-2 precursor FORM ICH-1S ICE8_HUMANISOQ9C0K4 Caspase-8 precursor FORM 7 ICEA_HUMANISO Q92851 Caspase-10precursor FORM B ICEA_HUMANISO Q92851 Caspase-10 precursor FORM CILF1_HUMANISO Q01167 Interleukin enhancer- FORM 2 binding factor 1IRF7_HUMANISO Q92985 Interferon regulatory factor 7 FORM C IRL1_HUMANISOQ01638 Interleukin 1 receptor-like FORM C 1 precursor ITP1_HUMANMAJORO14713 Integrin beta-1 binding ISOFORM protein 1 ITP1_HUMANISO O14713Integrin beta-1 binding FORM 2 protein 1 KC11_HUMANIS Q9HCP0 Caseinkinase I gamma 1 OFORM 1S isoform KLK2_HUMANIS P20151 Glandularkallikrein 2 OFORM 3 precursor KLKF_HUMANISO Q9H2R5 Kallikrein 15precursor FORM 2 LEF1_HUMANISO Q9UJU2 Lymphoid enhancer FORM B bindingfactor 1 LFA3_HUMANISO P19256 Lymphocyte function- FORM SHORT associatedantigen 3 precursor LIK1_HUMANISO P53667 LIM domain kinase 1 FORM 3LSHR_HUMANMAJOR P22888 Lutropin- ISOFORM choriogonadotropic hormonereceptor precursor LYST_HUMANMAJOR Q99698 Lysosomal trafficking ISOFORMregulator M2A2_HUMANISO P49641 Alpha-mannosidase IIx FORM SHORTMADI_HUMANISO O95405 Mothers against FORM 2 decapentaplegic homologinteracting protein MAP2_HUMAN P11137 Microtubule-associated MAJORprotein 2 ISOFORM MAP2_HUMANI P11137 Microtubule-associated SOFORM MAP2Cprotein 2 MAP4_HUMANI P27816 Microtubule-associated SOFORM 2 protein 4MAX_HUMANISO P25912 Max protein FORM 3 MK11_HUMANISO Q15759Mitogen-activated protein FORM BETA-2 kinase 11 MLH3_HUMANISO Q9UHC1 DNAmismatch repair FORM 2 protein Mlh3 MRP3_HUMANISO O15438 Canalicularmultispecific FORM 3A organic anion transporter 2 MRP3_HUMANISO O15438Canalicular multispecific FORM 3B organic anion transporter 2MSRE_HUMANISO P21757 Macrophage scavenger FORM II receptor types I andII MTF2_HUMANMAJOR Q9Y483 Metal-response element- ISOFORM bindingtranscription factor 2 NK31_HUMANMAJOR Q99801 Homeobox protein Nkx-31ISOFORM NXF5_HUMANMAJOR Q9H1B4 Nuclear RNA export factor 5 ISOFORMNXFS_HUMANISO Q9H1B4 Nuclear RNA export factor 5 FORM B NXFS_HUMANISOQ9H1B4 Nuclear RNA export factor 5 FORM C NXF5_HUMANISO Q9H1B4 NuclearRNA export factor 5 FORM D NXFS_HUMANISO Q9H1B4 Nuclear RNA exportfactor 5 FORM E PHMX_HUMANISO Q96QS1 Phemx protein FORM 4 PHMX_HUMANISOQ96QS1 Phemx protein FORM 5 PML_HUMANISO P29590 Probable transcriptionFORM PML-3B factor PML PPE1_HUMANISO O14829 Serine/threonine proteinFORM 2 phosphatase with EF- hands-1 PPT2_HUMANISO Q9UMR5Palmitoyl-protein FORM 2 thioesterase 2 precursor PRD7_HUMANMAJOR Q9NQW5PR-domain zinc finger ISOFORM protein 7 PSA7_HUMANISO O14818 Proteasomesubunit alpha FORM 4 type 7 PSD4_HUMANISO P55036 26S proteasome non-FORM RPN10E ATPase regulatory subunit 4 PSN1_HUMANISO P49768 Presenilin1 FORM I-374 PTPD_HUMANMAJOR P23468 Protein-tyrosine ISOFORM phosphatasedelta precursor R51D_HUMANISO O94908 DNA repair protein FORM 2 RAD51homolog 4 RBMS_HUMAN Q93062 RNA-binding protein with MAJOR multiplesplicing ISOFORM RED1_HUMANMAJOR P78563 Double-stranded RNA- ISOFORMspecific editase 1 RHD_HUMANMAJOR Q9UQ21 Blood group Rh ISOFORMRHD_HUMANISO Q9UQ21 Blood group Rh FORM 3 RYK_HUMANMAJOR P34925Tyrosine-protein kinase ISOFORM RYK precursor RYK_HUMANIS P34925Tyrosine-protein kinase OFORM 2 RYK precursor SCA1_ HUMANISO O15126Secretory carrier-associated FORM 2 membrane protein 1 SEN7_HUMANMAJORQ9BQF6 Sentrin-specific protease 7 ISOFORM SFR5_HUMANISO Q13243 Splicingfactor FORM SRP40-2 arginine/serine-rich 5 SHX2_HUMANMAJOR O60902 Shortstature homeobox ISOFORM protein 2 SNB2 HUMANISO Q13425Beta-2-syntrophin FORM 2 SNXD_HUMANISO Q9Y5W8 Sorting nexin 13 FORM 2SON_HUMANISO Q9UPY0 SON protein FORM C SON_HUMANISO Q9UPY0 SON proteinFORM E SUR5_HUMANISO Q15528 Surfeit locus protein 5 FORM SURF5AT10B_HUMANMAJOR O14763 Tumor necrosis factor ISOFORM receptorsuperfamily member 10B precursor T10B HUMANISO O14763 Tumor necrosisfactor FORM SHORT receptor superfamily member 10B precursorTM31_HUMANISO Q9BZY9 Tripartite motif protein 31 FORM BETA TNR6_HUMANISOP25445 Tumor necrosis factor FORM 2 receptor superfamily member 6precursor TNR6_HUMANISO P25445 Tumor necrosis factor FORM 3 receptorsuperfamily member 6 precursor TNR6_HUMANISO P25445 Tumor necrosisfactor FORM 4 receptor superfamily member 6 precursor TNR6_HUMANISOP25445 Tumor necrosis factor FORM 5 receptor superfamily member 6precursor TPA_HUMANISO P00750 Tissue-type plasminogen FORM SHORTactivator precursor TPO HUMANMAJOR P40225 Thrombopoietin precursorISOFORM TR12_HUMANISO Q99831 Tumor necrosis factor FORM 12 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 3 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 4 receptor superfamily member 12precursor TR12_HUMANISO Q99831 Tumor necrosis factor FORM 5 receptorsuperfamily member 12 precursor TR12_HUMANISO Q99831 Tumor necrosisfactor FORM 6 receptor superfamily member 12 precursor TR12_HUMANISOQ99831 Tumor necrosis factor FORM 7 receptor superfamily member 12precursor U7I3_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 2 enzyme 7interacting protein 3 U713_HUMANISO Q9BYM8 Ubiquitin conjugating FORM 4enzyme 7 interacting protein 3 USH3_HUMANISO P58418 Usher syndrome type3 FORM B protein WS14_HUMANISO Q9NP71 Williams-Beuren syndrome FORM 5chromosome region 14 protein XE7_HUMANISO Q02040 B-lymphocyte antigenFORM SHORT precursor Z236_HUMANISO Q9UL36 Zinc finger protein 236 FORM AZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM 1 ZAN_HUMANISO Q9BXN9Zonadhesin precursor FORM 2 ZAN_HUMANISO Q9BXN9 Zonadhesin precursorFORM 4 ZAN_HUMANISO Q9BXN9 Zonadhesin precursor FORM
 5.


8. The product of claim 7 wherein the dataset of target putative proteinisoform sequences comprises isoforms of Calpain-10, CLK1, CLK2, CLK3,and LARD/TNFRSF12.